ß-Tricalcium Phosphate-Modified Aerogel Containing PVA/Chitosan Hybrid Nanospun Scaffolds for Bone Regeneration.

Electrospinning has recently been recognized as a potential method for use in biomedical applications such as nanofiber-based drug delivery or tissue engineering scaffolds. The present study aimed to demonstrate the electrospinning preparation and suitability of ß-tricalcium phosphate-modified aerogel containing polyvinyl alcohol/chitosan fibrous meshes (BTCP-AE-FMs) for bone regeneration under in vitro and in vivo conditions. The mesh physicochemical properties included a 147 ± 50 nm fibrous structure, in aqueous media the contact angles were 64.1 ± 1.7°, and it released Ca, P, and Si. The viability of dental pulp stem cells on the BTCP-AE-FM was proven by an alamarBlue assay and with a scanning electron microscope. Critical-size calvarial defects in rats were performed as in vivo experiments to investigate the influence of meshes on bone regeneration. PET imaging using 18F-sodium fluoride standardized uptake values (SUVs) detected 7.40 ± 1.03 using polyvinyl alcohol/chitosan fibrous meshes (FMs) while 10.72 ± 1.11 with BTCP-AE-FMs after 6 months. New bone formations were confirmed by histological analysis. Despite a slight change in the morphology of the mesh because of cross-linking, the BTCP-AE-FM basically retained its fibrous, porous structure and hydrophilic and biocompatible character. Our experiments proved that hybrid nanospun scaffold composite mesh could be a new experimental bone substitute bioactive material in future medical practice.

Early-stage growth and elemental composition patterns of Brassica napus L. in response to Cd-Zn contamination.

Soil contamination caused by the presence of Cd and the excess amount of Zn is a widespread concern in agricultural areas, posing significant risks to the growth and development of crops. In this paper, the early-stage development and metal (Cd and Zn) accumulation potential of rapeseed (Brassica napus L.) grown under different metal application schemes were assessed by determining radicle and hypocotyl length and the micro- and macro elemental composition of plantlets after 24, 72, and 120 h. The results indicated that the single and co-application of Cd and Zn significantly reduced the radicle and hypocotyl lengths. Accumulation intensity for Cd and Zn was affected by Cd and the combination of Cd and Zn in the solution, respectively. In addition, both metals significantly influenced the tissue Mn and had a minor effect on Cu and Fe concentrations. Both Cd and Zn significantly affected macro element concentrations by decreasing tissue Ca and influencing K and Mg concentrations in a dose- and exposure time-dependent manner. These findings specify the short-term and support the long-term use of rapeseed in remediation processes. However, interactions of metals are crucial in determining the concentration patterns in tissues, which deserves more attention in future investigations.

Isotopic study of honey documents widespread plant uptake of old carbon in North America.

A comprehensive understanding of carbon cycling pathways in the soil-plant system is needed to develop models that accurately predict global carbon reservoir responses to anthropogenic perturbations. Honey is a carbon-rich natural food produced by wild and managed pollinating insects all over the world; the composition of a single sample is a function of millions of pollinator-plant interactions. We studied the 13C/12C and Δ14C of 121 honey samples sourced from the United States, and found a significant older carbon contribution. The effect is observed from 25 to 45° latitude, not correlated with 13C/12C, and consistent with a previously published study on European honeys. In specific cases, the measured values were up to 20 ‰ (Δ14C) higher than the expected atmospheric 14CO2 value for the given year, which shows a significant older carbon contribution. We hypothesize that the older carbon is from plant liquids derived in part from soil carbon or stored nonstructural carbohydrates from plants, which shifts the calibrated age of the sample by 5 years or more. Our work is the first to describe the widespread occurrence of older carbon in honey and shows that radiocarbon measurements can be a powerful tool to trace carbon allocations in terrestrial food webs and detect the atmosphere-soil-plant carbon cycle contributions.

Heavy metals in unrecorded Albanian rakia: A pilot study on a potential public health risk.

Unrecorded alcohol has been linked to illness above and beyond that caused by ethanol alone because of the presence of toxic contaminants. While it can be found in all countries, consumption is high in Albania, where it is frequently consumed as a fruit brandy known as rakia. Among the contaminants identified previously in such products, metals including lead have been detected at levels posing a risk to health but there is little information on their presence in rakia. To fill this gap, we measured the level of ethanol and 24 elements among them toxic metals in 30 Albanian rakia samples. We found that 63.3% of rakia samples had ethanol concentration above 40% v/v. We also showed that there was a significant difference between the measured [mean: 46.7% v/v, interquartile range (IQR): 43.4-52.1% v/v] and reported (mean: 18.9% v/v, IQR: 17.0-20.0% v/v) concentrations of ethanol in rakia. Among the metals detected, aluminium, copper, iron, manganese, lead, and zinc were present in rakia samples at concentrations ranging between 0.013 and 0.866 mg/l of pure alcohol (pa), 0.025-31.629 mg/l of pa, 0.004-1.173 mg/l of pa, 0.185-45.244 mg/l of pa, 0.044-1.337 mg/l of pa, and 0.004-10.156 mg/l of pa, respectively. Copper and lead were found to be the greatest concern posing a potential public health risk. Although the estimated daily intake of these heavy metals from unrecorded rakia was below their toxicological threshold, the concentrations of lead and copper exceeded their limit value of 0.2 and 2.0 mg/l of pa specified for spirits in 33% and 90% of samples, respectively. Therefore, the possibility of adverse health effects cannot be excluded completely. Our findings highlight the need for action by policymakers against the risks posed by these products in Albania.

Species- and Metal-Specific Responses of the Ionome of Three Duckweed Species under Chromate and Nickel Treatments.

In this study, growth and ionomic responses of three duckweed species were analyzed, namely Lemna minor, Landoltia punctata, and Spirodela polyrhiza, were exposed for short-term periods to hexavalent chromium or nickel under laboratory conditions. It was found that different duckweed species had distinct ionomic patterns that can change considerably due to metal treatments. The results also show that, because of the stress-induced increase in leaf mass-to-area ratio, the studied species showed different order of metal uptake efficiency if plant area was used as unit of reference instead of the traditional dry weight-based approach. Furthermore, this study revealed that µXRF is applicable in mapping elemental distributions in duckweed fronds. By using this method, we found that within-frond and within-colony compartmentation of metallic ions were strongly metal- and in part species-specific. Analysis of duckweed ionomics is a valuable approach in exploring factors that affect bioaccumulation of trace pollutants by these plants. Apart from remediating industrial effluents, this aspect will gain relevance in food and feed safety when duckweed biomass is produced for nutritional purposes.

Rape, sunflower and forest honeys for long-term environmental monitoring: Presence of indicator elements and non-photosynthetic carbon in old Hungarian samples.

In this paper, we present the time-dependent elemental composition and AMS radiocarbon dating results of 36 rape, sunflower and forest honey samples, collected between 1985 and 2018 in geographically close locations. Based on the elemental information, we conclude that bee products regardless the type provide useful environmental information of the previous decades, such as the decreasing trend of airborne Pb emission can be traced. However, radiocarbon results agree less with the atmospheric bomb peak. Random offsets were observed in the specific radiocarbon activity of the honey samples indicating that rape, sunflower and forest honey samples are not as reliable materials for radiocarbon dating as acacia honeys. The radiocarbon results show that the rape, sunflower and forest honey samples can contain non-photosynthetic carbon, presumably derived from the soil. Thus, the complex application of honey samples for environmental reconstruction requires the species-separated investigation of bee products to reveal their adaptability for assessment approaches.

Physiological and transcriptional profiling of surfactin exerted antifungal effect against Candida albicans.

Given the risk of Candida albicans overgrowth in the gut, novel complementary therapies should be developed to reduce fungal dominancy. This study highlights the antifungal characteristics of a Bacillus subtilis-derived secondary metabolite, surfactin with high potential against C. albicans. Surfactin inhibited the growth of C. albicans following a 1-hour exposure, in addition to reduced adhesion and morphogenesis. Specifically, surfactin did not affect the level of reactive oxygen species but increased the level of reduced glutathione. Surprisingly, ethanol production was increased following 2 h of surfactin exposure. Surfactin treatment caused a significant reduction in intracellular iron, manganese and zinc content compared to control cells, whereas the level of copper was not affected. Alongside these physiological properties, surfactin also enhanced fluconazole efficacy. To gain detailed insights into the surfactin-related effects on C. albicans, genome-wide gene transcription analysis was performed. Surfactin treatment resulted in 1390 differentially expressed genes according to total transcriptome sequencing (RNA-Seq). Of these, 773 and 617 genes with at least a 1.5-fold increase or decrease in transcription, respectively, were selected for detailed investigation. Several genes involved in morphogenesis or related to metabolism (e.g., glycolysis, ethanol and fatty acid biosynthesis) were down-regulated. Moreover, surfactin decreased the expression of ERG1, ERG3, ERG9, ERG10 and ERG11 involved in ergosterol synthesis, whereas genes associated with ribosome biogenesis and iron metabolism and drug transport-related genes were up-regulated. Our data demonstrate that surfactin significantly influences the physiology and gene transcription of C. albicans, and could contribute to the development of a novel innovative complementary therapy.

The Effect of Heat Treatment of ß-Tricalcium Phosphate-Containing Silica-Based Bioactive Aerogels on the Cellular Metabolism and Proliferation of MG63 Cells.

ß-Tricalcium phosphate was combined with silica aerogel in composites prepared using the sol-gel technique and supercritical drying. The materials were used in this study to check their biological activity and bone regeneration potential with MG63 cell experiments. The composites were sintered in 100 °C steps in the range of 500-1000 °C. Their mechanical properties, porosities, and solubility were determined as a function of sintering temperature. Dissolution studies revealed that the released Ca-/P molar ratios appeared to be in the optimal range to support bone tissue induction. Cell viability, ALP activity, and type I collagen gene expression results all suggested that the sintering of the compound at approximately 700-800 °C as a scaffold could be more powerful in vivo to facilitate bone formation within a bone defect, compared to that documented previously by our research team. We did not observe any detrimental effect on cell viability. Both the alkaline phosphatase enzyme activity and the type I collagen gene expression were significantly higher compared with the control and the other aerogels heat-treated at different temperatures. The mesoporous silica-based aerogel composites containing ß-tricalcium phosphate particles treated at temperatures lower than 1000 °C produced a positive effect on the osteoblastic activity of MG63 cells. An in vivo 6 month-long follow-up study of the mechanically strongest 1000 °C sample in rat calvaria experiments provided proof of a complete remodeling of the bone.

Changes in Lithium Levels in Bees and Their Products Following Anti-Varroa Treatment.

The biggest threat to beekeeping is varroosis caused by the mite Varroa destructor. Chemicals available to treat this fatal disease may present problems of resistance or inconsistent efficacy. Recently, lithium chloride has appeared as a potential alternative. To date, the amount of residue lithium treatments may leave in honeybee products is poorly understood. Honeybees were fed with 25 mM lithiated sugar syrup, which was used in earlier studies. The accumulation and elimination of the lithium were monitored in bees and their products for 22 days. Lithium concentration increased in the entire body of the bees to day 4 post-treatment and then recovered rapidly to the control level. Lithium exposure was found to affect uncapped honey in the short term (<16 days), but ripe (capped) honey measured at the end of the trial remained affected. On the other hand, lithium treatment left beeswax lithium-free. Based on these data, we propose that comprehensive research on harvested honey is needed to decide on the veterinary use of lithium.

Iron and Manganese Retention of Juvenile Zebrafish (Danio rerio) Exposed to Contaminated Dietary Zooplankton (Daphnia pulex)-a Model Experiment.

In present study the effect of iron (Fe) and manganese (Mn) contamination was assessed by modeling a freshwater food web of water, zooplankton (Daphnia pulex), and zebrafish (Danio rerio) under laboratory conditions. Metals were added to the rearing media of D. pulex, and enriched zooplankton was fed to zebrafish in a feeding trial. The elemental analysis of rearing water, zooplankton, and fish revealed significant difference in the treatments compared to the control. In D. pulex the Mn level increased almost in parallel with the dose of supplementation, as well as the Fe level differed statistically. A negative influence of the supplementation on the fish growth was observed: specific growth rate (SGR%) and weight gain (WG) decreased in Fe and Mn containing treatments. The redundancy analysis (RDA) of concentration data showed strong correlation between the rearing water and D. pulex, as well as the prey organism of Fe- and Mn-enriched D. pulex and the predator organism of D. rerio. The bioconcentration factors (BCF) calculated for water to zooplankton further proved the relationship between the Fe and Mn dosage applied in the treatments and measured in D. pulex. Trophic transfer factor (TTF) results also indicate that significant retention of the metals occurred in D. rerio individuals, however, in a much lower extent than in the water to zooplankton stage. Our study suggests that Fe and Mn significantly accumulate in the lower part of the trophic chain and retention is effective through the digestive track of zebrafish, yet no biomagnification occurs. Graphical abstract.

The Negative Effect of Protein Phosphatase Z1 Deletion on the Oxidative Stress Tolerance of Candida albicans Is Synergistic with Betamethasone Exposure.

The glucocorticoid betamethasone (BM) has potent anti-inflammatory and immunosuppressive effects; however, it increases the susceptibility of patients to superficial Candida infections. Previously we found that this disadvantageous side effect can be counteracted by menadione sodium bisulfite (MSB) induced oxidative stress treatment. The fungus specific protein phosphatase Z1 (CaPpz1) has a pivotal role in oxidative stress response of Candida albicans and was proposed as a potential antifungal drug target. The aim of this study was to investigate the combined effects of CaPPZ1 gene deletion and MSB treatment in BM pre-treated C. albicans cultures. We found that the combined treatment increased redox imbalance, enhanced the specific activities of antioxidant enzymes, and reduced the growth in cappz1 mutant (KO) strain. RNASeq data demonstrated that the presence of BM markedly elevated the number of differentially expressed genes in the MSB treated KO cultures. Accumulation of reactive oxygen species, increased iron content and fatty acid oxidation, as well as the inhibiting ergosterol biosynthesis and RNA metabolic processes explain, at least in part, the fungistatic effect caused by the combined stress exposure. We suggest that the synergism between MSB treatment and CaPpz1 inhibition could be considered in developing of a novel combinatorial antifungal strategy accompanying steroid therapy.

Transcriptional Profiling of the Candida auris Response to Exogenous Farnesol Exposure.

The antifungal resistance threat posed by Candida auris necessitates bold and innovative therapeutic options. Farnesol is a quorum-sensing molecule with a potential antifungal and/or adjuvant effect; it may be a promising candidate in alternative treatment regimens. To gain further insights into the farnesol-related effect on C. auris, genome-wide gene transcription analysis was performed using transcriptome sequencing (RNA-Seq). Farnesol exposure resulted in 1,766 differentially expressed genes. Of these genes, 447 and 304 genes with at least 1.5-fold increase or decrease in transcription, respectively, were selected for further investigation. Genes involved in morphogenesis, biofilm events (maturation and dispersion), gluconeogenesis, iron metabolism, and regulation of RNA biosynthesis showed downregulation, whereas those related to antioxidative defense, transmembrane transport, glyoxylate cycle, fatty acid ß-oxidation, and peroxisome processes were upregulated. In addition, farnesol treatment increased the transcription of certain efflux pump genes, including MDR1, CDR1, and CDR2. Growth, measured by the change in the number of CFU, was significantly inhibited within 2 h of the addition of farnesol (5.8 × 107 ± 1.1 × 107 and 1.1 × 107 ± 0.3 × 107 CFU/ml for untreated control and farnesol-exposed cells, respectively) (P < 0.001). In addition, farnesol treatment caused a significant reduction in intracellular iron (152.2 ± 21.1 versus 116.0 ± 10.0 mg/kg), manganese (67.9 ± 5.1 versus 18.6 ± 1.8 mg/kg), and zinc (787.8 ± 22.2 versus 245.8 ± 34.4 mg/kg) (P < 0.05 to 0.001) compared to untreated control cells, whereas the level of cooper was significantly increased (274.6 ± 15.7 versus 828.8 ± 106.4 mg/kg) (P < 0.001). Our data demonstrate that farnesol significantly influences the growth, intracellular metal ion contents, and gene transcription related to fatty acid metabolism, which could open new directions in developing alternative therapies against C. auris. IMPORTANCE Candida auris is a dangerous fungal pathogen that causes outbreaks in health care facilities, with infections associated with a high mortality rate. As conventional antifungal drugs have limited effects against the majority of clinical isolates, new and innovative therapies are urgently needed. Farnesol is a key regulator molecule of fungal morphogenesis, inducing phenotypic adaptations and influencing biofilm formation as well as virulence. Alongside these physiological modulations, it has a potent antifungal effect alone or in combination with traditional antifungals, especially at supraphysiological concentrations. However, our knowledge about the mechanisms underlying this antifungal effect against C. auris is limited. This study has demonstrated that farnesol enhances the oxidative stress and reduces the fungal survival strategies. Furthermore, it inhibits manganese, zinc transport, and iron metabolism as well as increases fungal intracellular copper content. In addition, metabolism was modulated toward ß-oxidation. These results provide definitive explanations for the observed antifungal effects.

Analysis of honey using capillary electrophoresis with electrokinetic injection - Direct analysis of viscous samples without sample pretreatment.

Although the electrokinetic injection mode is applicable for direct introduction of ionic components from highly viscous samples, this is the first work which studies this special feature of capillary electrophoresis for this purpose. 9 metal cations were determined in honey utilizing direct injection, zone electrophoretic separation and indirect UV detection of the sample components. Reproducibility better than 4 RSD% was found for peak areas and the LOD values ranged between 0.045 and 0.44 mg/kg when injection parameters were set to 7.5 kV × 5 s. In order to remedy the quantitation problems arising from mobility and matrix bias, the applicability of the internal universal calibration was examined. It was found that in cases where the mobilities of cations do not differ from the mobilities of the monitoring ion and the internal universal standard (IUS) by more than 10%, the obtained results agreed well with the data measured with ICP-OES.

Honey as an indicator of long-term environmental changes: MP-AES analysis coupled with 14C-based age determination of Hungarian honey samples.

Several studies show that the elemental content of honey entirely depends on the botanical and geographical origin, but the information is incomplete regarding its time-dependent composition changes. Twenty-six acacia and three honey samples with unknown botanical origin were collected between 1958 and 2018 and analysed for elemental composition by Microwave Plasma Atomic Emission Spectrometry (MP-AES). The elemental analysis was coupled with independent dating method by Accelerator Mass Spectrometry (AMS) to confirm the calendar age of the honey samples and test the possibility of radiocarbon based dating of bee products, which has not been applied before. According to the analytical measurements and statistical analysis, we can conclude that the elemental composition shows change with time in the acacia honey during the last five decades. We have proven that honey preserves carbon isotopic and elemental information of its production time and thus can be applied as an environmental indicator (e.g. trace urban pollutants, precipitation, local industrial or agricultural emission) in reconstruction studies by analysing the non-degradable mineral content. Our results further show that acacia honey is a suitable material for radiocarbon dating, proved by the results compared to the atmospheric radiocarbon bomb-peak. The new approach presented for investigation of honey by radiocarbon-based age determination coupled with elemental analysis can be used in biological, dietary, archaeological or other multidisciplinary studies as well. Some samples show slightly depleted radiocarbon content. This could be an indication of local fossil CO2 emission. Based on these depleted 14C results, honey could be used for atmospheric monitoring of fossil CO2 urban or industrial hot-spots.