A comprehensive study on Geranium robertianum L. antibacterial potential.

AIMS: The research aimed to optimize the ultrasound-assisted extraction of secondary metabolites and the antibacterial activity of the plant species Geranium robertianum. The phytochemical profiles of the optimized extracts, as well as their antibacterial and synergistic activity with an antibiotic and their potential mechanisms of action and cytotoxicity, were examined. METHODS AND RESULTS: Response Surface Methodology was used to optimize extraction conditions. Optimized ethanol and acetone extracts were tested via microdilution, checkerboard, time-kill kinetics, and cell membrane permeability methods. The extracts displayed broad antibacterial activity with minimum inhibitory concentrations ranging from 1.25 to 20 mg ml-1. In addition, the extract synergistically reacted with gentamicin against gentamicin-resistant strains of Escherichia coli and Staphylococcus aureus, enhancing the efficacy of the antibiotic up to 32-fold. The extracts demonstrated strain-dependent bactericidal activity in a 24-h time interval. They increase the permeability of the cell membrane, thus disrupting its normal functioning. The cytotoxic concentration (CC50) on human keratinocytes was 1771.24 ± 5.78 µg ml-1 for ethanol extract, and 958.01 ± 6.14 µg ml-1 for acetone extract. Kaempferol, ellagic acid, quercetin, and rutin were recognized as the main components in both extracts. CONCLUSIONS: The findings of this study indicate that the extracts of G. robertianum can be considered as potential natural antibacterial agents in the control of microorganisms.

Geranium robertianum L. tolerates various soil types burdened with heavy metals.

Many heavy metals (HMs) are essential micronutrients for the growth and development of plants. However, human activities such as mining, smelting, waste disposal, and industrial processes have led to toxic levels of HMs in soil. Fortunately, many plant species have developed incredible adaptive mechanisms to survive and thrive in such harsh environments. As a widespread and ruderal species, Geranium robertianum L. inhabits versatile soil types, both polluted and unpolluted. Considering the ubiquity of G. robertianum, the study aimed to determine whether geographically distant populations can tolerate HMs. We collected soil and plant samples from serpentine, an anthropogenic heavy metal contaminated, and a non-metalliferous site to study the physiological state of G. robertianum. HMs in soil and plants were determined using flame atomic absorption spectrometry. Spectrophotometric methods were used to measure the total content of chlorophylls a and b, total phenolics, phenolic acids, flavonoids, and proline. Principal component analysis (PCA) was used to investigate the potential correlation between HMs concentrations gathered from various soil types and plant samples and biochemical data acquired for plant material. A statistically significant difference was observed for all localities regarding secondary metabolite parameters. A positive correlation between Ni and Zn in soil and Ni and Zn in plant matter was observed (p<0.0005) indicating higher absorption. Regardless of high concentrations of heavy metals in investigated soils, G. robertianum displayed resilience and was capable of thriving. These results may be ascribed to several protective mechanisms that allow G. robertianum to express normal growth and development and act as a pioneer species.

Herb Robert's Gift against Human Diseases: Anticancer and Antimicrobial Activity of Geranium robertianum L.

Geranium robertianum L. is a widely distributed plant used as a traditional herbal medicine, but the knowledge of its biological properties still needs to be improved. Thus, the purpose of this presented research was to assess the phytochemical profile of extracts from aerial parts of G. robertianum, commercially available in Poland and to study their anticancer potential and antimicrobial properties, including the antiviral, antibacterial, and antifungal effects. Additionally, the bioactivity of fractions obtained from the hexane and ethyl acetate extract was analyzed. The phytochemical analysis revealed the presence of organic and phenolic acids, hydrolysable tannins (gallo- and ellagitannins), and flavonoids. Significant anticancer activity was found for G. robertianum hexane extract (GrH) and ethyl acetate extract (GrEA) with an SI (selectivity index) between 2.02 and 4.39. GrH and GrEA inhibited the development of HHV-1-induced cytopathic effect (CPE) in virus-infected cells and decreased the viral load by 0.52 log and 1.42 log, respectively. Among the analyzed fractions, only those obtained from GrEA showed the ability to decrease the CPE and reduce the viral load. The extracts and fractions from G. robertianum showed a versatile effect on the panel of bacteria and fungi. The highest activity was observed for fraction GrEA4 against Gram-positive bacteria, including Micrococcus luteus ATCC 10240 (MIC 8 µg/mL), Staphylococcus epidermidis ATCC 12228 (MIC 16 µg/mL), Staphylococcus aureus ATCC 43300 (MIC 125 µg/mL), Enterococcus faecalis ATCC 29212 (MIC 125 µg/mL), and Bacillus subtilis ATCC 6633 (MIC 125 µg/mL). The observed antibacterial effect may justify the traditional use of G. robertianum to treat hard-to-heal wounds.

Evidence of spontaneous selfing and disomic inheritance in Geranium robertianum.

Knowing species' breeding system and mating processes occurring in populations is important not only for understanding population dynamics, gene flow processes, and species' response to climate change, but also for designing control plans of invasive species. Geranium robertianum, a widespread biennial herbaceous species showing high morphological variation and wide ecological amplitude, can become invasive outside its distribution range. A mixed-mating system may be expected given the species' floral traits. However, autonomous selfing is considered as a common feature. Genetic variation and structure, and so population mating processes, have not been investigated in wild populations. We developed 15 polymorphic microsatellite markers to quantify genetic variation and structure in G. robertianum. To investigate whether selfing might be the main mating process in natural conditions, we sampled three generations of plants (adult, F1, and F2) for populations from the UK, Spain, Belgium, Germany, and Sweden, and compared open-pollinated with outcrossed hand-pollinated F2 progeny. The highly positive Wright's inbreeding coefficient (F IS) values in adults, F1, and open-pollinated F2 progeny and the low F IS values in outcross F2 progeny supported autonomous selfing as the main mating process for G. robertianum in wild conditions, despite the presence of attractive signals for insect pollination. Genetic differentiation among samples was found, showing some western-eastern longitudinal trend. Long-distance seed dispersal might have contributed to the low geographic structure. Local genetic differentiation may have resulted not only from genetic drift effects favored by spontaneous selfing, but also from ecological adaptation. The presence of duplicate loci with disomic inheritance is consistent with the hypothesis of allotetraploid origin of G. robertianum. The fact that most microsatellite markers behave as diploid loci with no evidence of duplication supports the hypothesis of ancient polyploidization. The differences in locus duplication and the relatively high genetic diversity across G. robertianum range despite spontaneous autonomous selfing suggest multiple events of polyploidization.

Survival on railway tracks of Geranium robertianum-a glyphosate-tolerant plant.

Geranium robertianum is a herbaceous plant that prefers shady and fertile forest habitats. However, it also occurs on railway tracks, where there are difficult conditions for plant growth and regular herbicide spraying (in high concentrations, twice a year). One of the most commonly used herbicides in railway areas is glyphosate. The effect of the glyphosate on the G. robertianum plants found on railway tracks and in nearby forests in north-eastern Poland was checked. The aim of the study was to explain how G. robertianum can survive on railway tracks despite spraying with the glyphosate. Increased tolerance to the glyphosate of the G. robertianum plants from track populations was demonstrated compared to the plants from forest populations that had not previously been in contact with the herbicide. After 35 days after treatment with the herbicide, 75% of the plants from the observed forest populations withered, while only 38% did from the track populations. Ultrastructure of plant leaf cells from forest populations was strongly disturbed, which was not observed in plants from track populations. It was also shown that plants from track populations accumulated more glyphosate and AMPA in their tissues than plants from forest populations. The obtained results indicate that long-term use of herbicides may cause formation of biotypes of plants resistant to a given herbicide. This fact explains the possibility of G. robertianum occurring on railway tracks, despite spraying with the glyphosate. It is also a manifestation of microevolutionary processes.

Arbuscular mycorrhizal fungi change host plant DNA methylation systemically.

DNA methylation is an important epigenetic mechanism regulating gene expression in plants. DNA methylation has been shown to vary among species and also among plant tissues. However, no study has evaluated whether arbuscular mycorrhizal (AM) fungi affect DNA methylation levels in a tissue-specific manner. We investigated whether symbiosis with AM fungi affects DNA methylation in the host, focusing on different plant tissues (roots versus leaves) and across time. We carried out a 6-month pot experiment using Geranium robertianum in symbiosis with the AM fungus Funneliformis mosseae. Our results show that the pattern of total DNA methylation differed between leaves and roots and was related to when plants were harvested, confirming that DNA methylation is a process that occurs dynamically throughout an organism's lifetime. More importantly, the presence of AM fungus in roots of our experimental plants had a positive effect on total DNA methylation in both tissues. This study shows that colonisation by AM fungi can affect DNA methylation levels in their hosts and that plant DNA methylation varies in an age- and tissue-specific manner.

Investigation of the effectiveness of Syzygium aromaticum, Lavandula angustifolia and Geranium robertianum essential oils in the treatment of acute external otitis: a comparative trial with ciprofloxacin.

BACKGROUND: Antibiotics and anti-inflammatory agents are the mainstay of acute external otitis (AEO) treatment. The present study investigated the effectiveness of a combination herbal drop (Lamigex) composed of essential oils from Syzygium aromaticum, Lavandula angustifolia, and Geranium robertianum in the alleviation of AEO symptoms and compared its effects to those of ciprofloxacin 0.3% drop. METHODS: Seventy patients were randomly assigned to receive ciprofloxacin 0.3% (n = 35) or Lamigex (n = 35) drop. Each group was administered with three drops every 12 hours for a week. Patients were examined for AEO symptoms and ear discharge cultures at baseline as well as at the end of trial. Pain severity was also recorded using a visual analogue scale at baseline, the 3(rd) day, and the 7(th) day of the trial. RESULTS: All assessed symptoms (tenderness, itching, erythema, edema and discharge) were equally improved in the ciprofloxacin and Lamigex groups by the end of trial (p > 0.05). There were remarkable reductions in the visual analogue scale score by the end of trial in both groups (p < 0.001). However, the rate of pain improvement was not found to be significantly different between the groups, either at the 3(rd) or 7(th) day of trial (p > 0.05). The numbers of positive cultures for all tested microorganisms were clearly reduced by the end of the trial in both groups but were not significantly different between the groups (p > 0.05). CONCLUSION: The herbal combination drop that was investigated in the present study exhibited good efficacy in reducing the burden of infection as well as AEO symptoms.