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Species of the genus Drosera, known for carnivorous plants, such as sundew, have been traditionally used for centuries as medicinal plants. Efficacy-determining compounds are naphthoquinones and flavonoids. Flavonoids possess a broad spectrum of bioactive properties, including biofilm inhibitory activity. Biofilms render antibiotics ineffective, contributing to the current rise in antimicrobial resistance. In this study, the biofilm inhibitory activity of two European sundew species (Drosera rotundifolia and Drosera intermedia) grown agriculturally in Germany and four commercial sundew products (declared as Drosera longifolia, Drosera sp. and Drosera planta trit.) against three multidrug-resistant Escherichia coli strains was tested. The aim of the study was to comparatively investigate the biofilm inhibitory potential of sundew species extracts grown locally in northern Germany and commercial sundew products. The minimum biofilm inhibitory concentration of the European sundew species was approx. 35 µg mL−1. In comparison, commercial sundew products ranged in concentration from 75 to 140 µg mL−1. Additionally, individual compounds isolated from European sundew were tested. Among these compounds, biofilm inhibitory activity was determined for four of the eight substances, with 2″-O-galloyl hyperoside standing out for its activity (38 µg mL−1). The whole plant extracts of Drosera rotundifolia and Drosera intermedia proved to be more effective than the commercial products and the single compounds in its biofilm inhibition activity against Escherichia coli strains. Sundew extracts may serve as a potential therapeutic approach for targeting biofilm production.
The goal of this study was to assess the anticancer efficacy of chlorojanerin against various cancer cells. The effects of chlorojanerin on cell cytotoxicity, cell cycle arrest, and cell apoptosis were examined using MTT assay, propidium iodide staining, and FITC Annexin V assay. RT-PCR was employed to determine the expression levels of apoptosis-related genes. Furthermore, docking simulations were utilized to further elucidate the binding preferences of chlorojanerin with Bcl-2. According to MTT assay, chlorojanerin inhibited the proliferation of all tested cells in a dose-dependent manner with a promising effect against A549 lung cancer cells with an IC50 of 10 µM. Cell growth inhibition by chlorojanerin was linked with G2/M phase cell cycle arrest in A549 treated cells. Flow cytometry analysis indicated that the proliferation inhibition effect of chlorojanerin was associated with apoptosis induction in A549 cells. Remarkably, chlorojanerin altered the expression of many genes involved in apoptosis initiation. Moreover, we determined that chlorojanerin fit into the active site of Bcl-2 according to the molecular docking study. Collectively, our results demonstrate that chlorojanerin mediated an anticancer effect involving cell cycle arrest and apoptotic cell death and, therefore, could potentially serve as a therapeutic agent in lung cancer treatment.
In the search for alternative treatment options for infections with multi-resistant germs, traditionally used medicinal plants are currently being examined more intensively. In this study, the antimicrobial and anti-biofilm activities of 14 herbal drugs were investigated. Nine of the tested drugs were traditionally used in Europe for treatment of local infections. For comparison, another five drugs monographed in the European Pharmacopoeia were used. Additionally, the total tannin and flavonoid contents of all tested drugs were analyzed. HPLC fingerprints were recorded to obtain further insights into the components of the extracts. The aim of the study was to identify herbal drugs that might be useable for treatment of infectious diseases, even with multidrug resistant E. coli, and to correlate the antimicrobial activity with the total content of tannins and flavonoids. The agar diffusion test and anti-biofilm assay were used to evaluate the antimicrobial potential of different extracts from the plants. Colorimetric methods (from European Pharmacopeia) were used for determination of total tannins and flavonoids. The direct antimicrobial activity of most of the tested extracts was low to moderate. The anti-biofilm activity was found to be down to 10 µg mL−1 for some extracts. Tannin contents between 2.2% and 10.4% of dry weight and total flavonoid contents between 0.1% and 1.6% were found. Correlation analysis indicates that the antimicrobial and the anti-biofilm activity is significantly (p < 0.05) dependent on tannin content, but not on flavonoid content. The data analysis revealed that tannin-rich herbal drugs inhibit pathogens in different ways. Thus, some of the tested herbal drugs might be useable for local infections with multi-resistant biofilm-forming pathogens. For some of the tested drugs, this is the first report about anti-biofilm activity, as well as total tannin and flavonoid content.
This communication introduces the first-time application of high-resolution continuum-source molecular absorption spectrometry (HR CS MAS) for the quantification of a peptide. The graphite furnace technique was employed and the tripeptide glutathione (GSH) served as a model compound. Based on measuring sulfur in terms of carbon monosulfide (CS), a method was elaborated to analyze aqueous solutions of GSH. The most prominent wavelength of the CS molecule occurred at 258.0560 nm and was adduced for monitoring. The methodological development covered the optimization of the pyrolysis and vaporization temperatures. These were found optimally to be 250 °C and 2250 °C, respectively. Moreover, the effect of modifiers (zirconium, calcium, magnesium, palladium) on the absorption signals was investigated. The best results were obtained after permanent coating of the graphite tube with zirconium (total amount of 400 μg) and adding a combination of palladium (10 µL, 10 g L−1) and calcium (2 µL, 1 g L−1) as a chemical modifier to the probes (10 µL). Aqueous standard samples of GSH were used for the calibration. It showed a linear range of 2.5–100 µg mL−1 sulfur contained in GSH with a correlation coefficient R2 > 0.997. The developed method exhibited a limit of detection (LOD) and quantification (LOQ) of 2.1 µg mL−1 and 4.3 µg mL−1 sulfur, respectively. The characteristic mass accounted for 5.9 ng sulfur. The method confirmed the general suitability of MAS for the analysis of an oligopeptide. Thus, this study serves as groundwork for further development in order to extend the application of classical atomic absorption spectrometry (AAS).
Background
In addition to the broad dissemination of pathogenic extended-spectrum beta-lactamase (ESBL)-producing Escherichia (E.) coli in human and veterinary medicine and the community, their occurrence in wildlife and the environment is a growing concern. Wild birds in particular often carry clinically relevant ESBL-producing E. coli.
Objectives
We analyzed ESBL-producing and non-ESBL-producing E. coli obtained from wild birds in Mongolia to identify phylogenetic and functional characteristics that would explain the predominance of a particular E. coli clonal lineage in this area.
Methods
We investigated ESBL-producing E. coli using whole-genome sequencing and phylogenetics to describe the population structure, resistance and virulence features and performed phenotypic experiments like biofilm formation and adhesion to epithelial cells. We compared the phenotypic characteristics to non-ESBL-producing E. coli from the same background (Mongolian wild birds) and genomic results to publicly available genomes.
Results and Conclusion
We found ESBL-producing E. coli sequence type (ST) 1159 among wild birds in Mongolia. This clonal lineage carried virulence features typical for extra-intestinal pathogenic or enterotoxigenic E. coli. Comparative functional experiments suggested no burden of resistance in the ST1159 isolates, which is despite their carriage of ESBL-plasmids. Wild birds will likely disseminate these antibiotic-resistant pathogens further during migration.
Urbanization, industrialization, and intensification of agriculture have led to considerable heavy metal pollution across the globe, harming our ecosystems. Concentrations of arsenic (As), cadmium (Cd), copper (Cu), and lead (Pb) have been analysed in 249 eggshells collected between 2006 and 2021 from 83 female Common Cranes (Grus grus) nesting within north-eastern Germany. Information on the presence of trace elements in cranes from Europe and their potential adverse effects on the reproduction are largely missing. Only Cu and Pb were found to be present in eggshell samples. Levels of both metals did not exceed concentrations considered potentially toxic in birds and unhatched eggs did not contain higher metal concentrations compared to eggshell residues from hatched eggs. Statistical analysis revealed that trace element concentrations decreased significantly over the course of the study period. The ban of leaded gasoline in the early twenty-first century and strict limitations of heavy metal-based biocontrol products are likely responsible for this decrease over the years. However, as Cu levels gradually increase with increasing proportions of agricultural areas within the cranes’ home ranges, we suggest that considerable amounts of Cu originating from agricultural practises are still being released into the environment. We found no increase in metal concentrations in eggshells with increasing female age, suggesting that heavy metals do not accumulate in the circulatory systems of the adults over time. This study is the first to assess heavy metal contamination in Common Cranes and indicates the suitability of crane’s eggshells as bioindicator for monitoring environmental pollution.
Multidrug-resistant (MDR) Enterobacterales, including extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae, not only emerge in healthcare settings but also in other habitats, such as livestock and wildlife. The spread of these pathogens, which often combine resistance with high-level virulence, is a growing problem, as infections have become increasingly difficult to treat. Here, we investigated the occurrence of ESBL-producing E. coli and K. pneumoniae in fecal samples from two black-headed gull colonies breeding on two nature conservation islands in Western Pomerania, Germany. In addition to cloacal samples from adult birds (n = 211) and their nestlings (n = 99) during the 2021 breeding season, collective fecal samples (n = 29) were obtained. All samples were screened for ESBL producers, which were then subjected to whole-genome sequencing. We found a total of 12 ESBL-producing E. coli and K. pneumoniae consisting of 11 E. coli and 1 K. pneumoniae, and including the international high-risk E. coli sequence types (ST)131, ST38, and ST58. Eight of the investigated strains had a MDR genotype and carried a large repertoire of virulence-associated genes, including the pap operon, which is important for urinary tract infections. In addition, we identified many genes associated with adherence, biofilm formation, iron uptake, and toxin production. Finally, our analysis revealed the close phylogenetic relationship of ST38 strains with genomes originating from human sources, underlining their zoonotic and pathogenic character. This study highlights the importance of the One Health approach, and thus the interdependence between human and animal health and their surrounding environment.
Multidrug-resistant gram-negative pathogens such as Escherichia coli have become increasingly difficult to treat and therefore alternative treatment options are needed. Targeting virulence factors like biofilm formation could be one such option. Inhibition of biofilm-related structures like curli and cellulose formation in E. coli has been shown for different phenolic natural compounds like epigallocatechin gallate. This study demonstrates this effect for other structurally unrelated phenolics, namely octyl gallate, scutellarein and wedelolactone. To verify whether these structurally different compounds influence identical pathways of biofilm formation in E. coli a broad comparative RNA-sequencing approach was chosen with additional RT-qPCR to gain initial insights into the pathways affected at the transcriptomic level. Bioinformatical analysis of the RNA-Seq data was performed using DESeq2, BioCyc and KEGG Mapper. The comparative bioinformatics analysis on the pathways revealed that, irrespective of their structure, all compounds mainly influenced similar biological processes. These pathways included bacterial motility, chemotaxis, biofilm formation as well as metabolic processes like arginine biosynthesis and tricarboxylic acid cycle. Overall, this work provides the first insights into the potential mechanisms of action of novel phenolic biofilm inhibitors and highlights the complex regulatory processes of biofilm formation in E. coli.
The study aimed to examine the influence of a rotating magnetic field (RMF) of two different frequencies (5 and 50 Hz) on the expression of regulatory (agrA, hld, rot) and staphylococcal enterotoxin (SE—sea, sec, sel) genes as well as the production of SEs (SEA, SEC, SEL) by the Staphylococcus aureus FRI913 strain cultured on a medium supplemented with a subinhibitory concentration of trans-anethole (TA). Furthermore, a theoretical model of interactions between the bacterial medium and bacterial cells exposed to RMF was proposed. Gene expression and SEs production were measured using quantitative real-time PCR and ELISA techniques, respectively. Based on the obtained results, it was found that there were no significant differences in the expression of regulatory and SE genes in bacteria simultaneously cultured on a medium supplemented with TA and exposed to RMF at the same time in comparison to the control (unexposed to TA and RMF). In contrast, when the bacteria were cultured on a medium supplemented with TA but were not exposed to RMF or when they were exposed to RMF of 50 Hz (but not to TA), a significant increase in agrA and sea transcripts as compared to the unexposed control was found. Moreover, the decreased level of sec transcripts in bacteria cultured without TA but exposed to RMF of 50 Hz was also revealed. In turn, a significant increase in SEA and decrease in SEC and SEL production was observed in bacteria cultured on a medium supplemented with TA and simultaneously exposed to RMFs. It can be concluded, that depending on SE and regulatory genes expression as well as production of SEs, the effect exerted by the RMF and TA may be positive (i.e., manifests as the increase in SEs and/or regulatory gene expression of SEs production) or negative (i.e., manifests as the reduction in both aforementioned features) or none.
Abstract
Aim
To verify synergistic effects, we investigated the antimicrobial activity of seven phenolic phytochemicals (gallic acid; epicatechin; epigallocatechin gallate; daidzein; genistein; myricetin; 3‐hydroxy‐6‐methoxyflavone) in combination with six antibiotics against multidrug‐resistant isolates from the ESKAPE group.
Methods and Results
To investigate single phytochemicals and combinations, initial microdilution and checkerboard assays were used, followed by time‐kill assays to evaluate the obtained results. The research revealed that phenolic compounds on their own resulted in little or no inhibitory effects. During preliminary tests, most of the combinations resulted in indifference (134 [71.3%]). In all, 30 combinations led to antagonism (15.9%); however, 24 showed synergistic effects (12.8%). The main tests resulted in nine synergistic combinations for the treatment of four different bacteria strains, including two substances (3‐hydroxy‐6‐methoxyflavone, genistein) never tested before in such setup. Time‐kill curves for combinations with possible synergistic effects confirmed the results against Acinetobacter baumannii as the one with the greatest need for research.
Conclusions
The results highlight the potential use of antibiotic–phytocompound combinations for combating infections with multi‐resistant pathogens. Synergistic combinations could downregulate the resistance mechanisms of bacteria.
Significance and Impact of the Study
The aim of this study is to demonstrate the potential use of phenolic natural compounds in combination with conventional antibiotics against multidrug‐resistant bacteria of the ESKAPE group. Due to synergistic effects of natural phenolic compounds combined with antibiotics, pathogens that are already resistant to antibiotics could be resensitized as we were able to reduce their MICs back to sensitive. In addition, combination therapies could prevent the development of resistance by reducing the dose of antibiotics. This approach opens up the basis for future development of antimicrobial therapy strategies, which are so urgently needed in the age of multidrug‐resistant pathogens.