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Clays attributed to have medicinal properties have been used since prehistoric times and are still used today as complementary medicines, which has given rise to unregulated "bioceutical" clays to treat skin conditions. Recently, clays with antibacterial characteristics have been proposed as alternatives to antibiotics, potentially overcoming modern day antibiotic resistance. Clays with suggested antibacterial properties were examined to establish their effects on common wound-infecting bacteria. Geochemical, microscopical, and toxicological characterization of clay particulates, their suspensions and filtered leachates was performed on THP-1 and HaCaT cell lines. Cytoskeletal toxicity, cell proliferation/viability (MTT assays), and migration (scratch wounds) were further evaluated. Clays were assayed for antibacterial efficacy using minimum inhibitory concentration assays. All clays possessed a mineral content with antibacterial potential; however, clay leachates contained insufficient ions to have any antibacterial effects. All clay leachates displayed toxicity towards THP-1 monocytes, while clay suspensions showed less toxicity, suggesting immunogenicity. Reduced clay cytotoxicity on HaCaTs was shown, as many leachates stimulated wound-healing responses. The "Green" clay exhibited antibacterial effects and only in suspension, which was lost upon neutralization. pH and its interaction with clay particle surface charge is more significant than previously understood to emphasize dangers of unregulated marketing and unsubstantiated bioceutical claims.

Objective: A four-arm randomized controlled trial (RCT) conducted in Victoria, Australia, previously evaluated parent-report outcomes following Confident Body, Confident Child: a program for parents to promote healthful eating patterns and positive body image in pre-schoolers. This exploratory study evaluated data from children of parents in the trial at 18 months follow-up. Method: Participants were 89 children (58 girls, 31 boys) of parents across all RCT arms (group A: Confident Body, Confident Child (CBCC) resource + workshop, n = 27; group B: CBCC resource only, n = 26; group C: nutrition booklet, n = 18; group D: wait-list control, n = 18). Children's eating patterns, body image and weight bias were assessed via play-based interview. Results: Children of CBCC parents reported higher body esteem. Children of nutrition booklet parents reported stronger weight bias. Children of CBCC workshop parents reported lower External Eating. Discussion: This exploratory study suggests that CBCC may promote healthy eating patterns and child body image 18 months after parents receive the intervention.

The Southern Hemisphere family Casuarinaceae has a long fossil record, both macrofossils and pollen, none of which provides any evidence about the morphology of the precursor to the family. However, it has long been considered, from both molecular phylogenies and morphological data, that the extant genus Gymnostoma retains key ancestral states and the highly reduced leaf area is a result of a scleromorphic response to low soil nutrients. Gymnostoma has by far the earliest, most extensive and best preserved macrofossil record, beginning in the Late Paleocene. Modification of the stomatal location from superficial in Gymnostoma to encrypted in furrows in the other genera assisted in water conservation as species evolved. We conclude that the morphology of the living and fossil vegetative branchlets provides evidence that low soil nutrients (especially phosphorus) and high water availability in a relatively light limited environment were the original drivers for evolution in the Casuarinaceae. Reducing water availability (xeromorphy) in progressively higher light environments were the major drivers of post-Eocene evolution in this unique plant family.

© 2019 Stomatal traits have been shown to vary in predictable ways in response to environmental change in many plant species. As a consequence, stomatal traits in fossil leaves are sometimes used as proxies for past CO2 and climate. Here we investigate the influence of temperature, rainfall and CO2 on stomatal traits in plant cuticle fine details of Melaleuca quinquenervia. We use both modern and fossil leaves deposited over the last c. 7500 years to evaluate the effect of CO2, and modern leaves for climate variables. We found a significant negative relationship between stomatal size and density across both modern and fossil leaves of M. quinquenervia. However, we were unable to find any relationship between stomatal traits and CO2 across a range from 260 to 380 ppm. We were unable to find any robust relationships between stomatal traits and either evaporation or temperature using the modern dataset. Apogeotropic roots account for the lack of stomatal anatomy response to evaporation in sites that experiences inundation. We conclude that stomatal size is a highly plastic trait in this species and changes do not necessarily reflect functional changes in the leaves.

Premise of research. This study compares the pinnule morphology of three fossil Bowenia species, one Eobowenia (fossil) species, and several Bowenia fossil pinnule fragments with the two extant Bowenia species. Methodology. Pinnule area, stomatal density, and size have been measured on fossil and extant specimens. Measures have been correlated with solar radiation and temperature to ascertain any correlations. Environment and climate variables have been chosen, as they have changed since the Eocene and Lower Cretaceous when the Australian and South American fossils were growing. Pivotal results. Two of the fossil Bowenia species and Eobowenia have significantly smaller pinnules than the living species, and all the fossils have relatively small and sparse stomata compared with the living species. Extant Bowenia pinnule area is positively correlated with daily radiation and temperature. Conclusions. Those fossil species with small pinnules likely lived in relatively dark environments where small pinnules aided the capture of sunflecks and/or facilitated light capture. The smaller and sparser stomata may have limited the potential sites for fungal invasion in the extremely humid growth conditions without hampering overall growth rates in the prevailing very high atmospheric CO 2 levels.

© 2019 Stomatal traits have been shown to vary in predictable ways in response to environmental change in many plant species. As a consequence, stomatal traits in fossil leaves are sometimes used as proxies for past CO2 and climate. Here we investigate the influence of temperature, rainfall and CO2 on stomatal traits in plant cuticle fine details of Melaleuca quinquenervia. We use both modern and fossil leaves deposited over the last c. 7500 years to evaluate the effect of CO2, and modern leaves for climate variables. We found a significant negative relationship between stomatal size and density across both modern and fossil leaves of M. quinquenervia. However, we were unable to find any relationship between stomatal traits and CO2 across a range from 260 to 380 ppm. We were unable to find any robust relationships between stomatal traits and either evaporation or temperature using the modern dataset. Apogeotropic roots account for the lack of stomatal anatomy response to evaporation in sites that experiences inundation. We conclude that stomatal size is a highly plastic trait in this species and changes do not necessarily reflect functional changes in the leaves.

Premise of research. This study compares the pinnule morphology of three fossil Bowenia species, one Eobowenia (fossil) species, and several Bowenia fossil pinnule fragments with the two extant Bowenia species. Methodology. Pinnule area, stomatal density, and size have been measured on fossil and extant specimens. Measures have been correlated with solar radiation and temperature to ascertain any correlations. Environment and climate variables have been chosen, as they have changed since the Eocene and Lower Cretaceous when the Australian and South American fossils were growing. Pivotal results. Two of the fossil Bowenia species and Eobowenia have significantly smaller pinnules than the living species, and all the fossils have relatively small and sparse stomata compared with the living species. Extant Bowenia pinnule area is positively correlated with daily radiation and temperature. Conclusions. Those fossil species with small pinnules likely lived in relatively dark environments where small pinnules aided the capture of sunflecks and/or facilitated light capture. The smaller and sparser stomata may have limited the potential sites for fungal invasion in the extremely humid growth conditions without hampering overall growth rates in the prevailing very high atmospheric CO 2 levels.

Objective: To use in vitro biofilm models of wound bacterial isolates and compare the biofilms produced for different combinations of wound bacterial species. Method: In vitro biofilms, generated by Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus oralis and Micrococcus luteus in microtitre plates and a constant depth film fermentor (cDFF), were studied. The tested isolates all originated from chronic venous leg ulcers. biofilms of individual and dual combinations of these species were generated in microtitre plate wells at 37°c for 24–96 hours and also in the CDFF for 7 days. The extent of biofilm formation from these systems was then measured using crystal violet staining and/or total viable counts. Results: All the chronic wound bacteria formed biofilms (both individually and in mixed culture) in these models. In mixed species microtitre plate biofilms, both P. aeruginosa and S. aureus appeared to antagonise biofilm formation by S. oralis and M. luteus, with P. aeruginosa completely inhibiting the growth of these organisms. Similar effects were evident in the CDFF model, when all four bacterial species were added simultaneously, with M. luteus being ‘out-competed’ by the other organisms present and occurring at numbers at the limits of detection; however, there was an apparent increase in the numbers of S. oralis compared with its single culture equivalent. Conclusion: The study highlighted differences in biofilm formation ability for the tested species in both closed and open model systems. Using dual species biofilms, distinct species antagonism was observed with apparent antagonism of pathogenic species over ‘commensal’ ones. Such a finding provides insight into possible bacterial interactions during development of ‘non-healing’ wound biofilms.

Biofilms provide a reservoir of potentially infectious micro-organisms that are resistant to antimicrobial agents, and their importance in the failure of medical devices and chronic inflammatory conditions is increasingly being recognized. Particular research interest exists in the association of biofilms with wound infection and non-healing, i.e. chronic wounds. In this study, fluorescent in situ hybridization (FISH) was used in combination with confocal laser scanning microscopy (CLSM) to detect and characterize the spatial distribution of biofilm-forming bacteria which predominate within human chronic skin wounds (Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus sp. and Micrococcus sp.). In vitro biofilms were prepared using a constant-depth film fermenter and a reconstituted human epidermis model. In vivo biofilms were also studied using biopsy samples from non-infected chronic venous leg ulcers. The specificity of peptide nucleic acid (PNA) probes for the target organisms was confirmed using mixed preparations of planktonic bacteria and multiplex PNA probing. Identification and location of individual bacterial species within multi-species biofilms demonstrated that P. aeruginosa was predominant. CLSM revealed clustering of individual species within mixed-species biofilms. FISH analysis of archive chronic wound biopsy sections showed bacterial presence and allowed bacterial load to be determined. The application of this standardized procedure makes available an assay for identification of single- or multi-species bacterial populations in tissue biopsies. The technique provides a reliable tool to study bacterial biofilm formation and offers an approach to assess targeted biofilm disruption strategies in vivo.

Biofilms provide a reservoir of potentially infectious micro-organisms that are resistant to antimicrobial agents, and their importance in the failure of medical devices and chronic inflammatory conditions is increasingly being recognized. Particular research interest exists in the association of biofilms with wound infection and non-healing, i.e. chronic wounds. In this study, fluorescent in situ hybridization (FISH) was used in combination with confocal laser scanning microscopy (CLSM) to detect and characterize the spatial distribution of biofilm-forming bacteria which predominate within human chronic skin wounds (Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus sp. and Micrococcus sp.). In vitro biofilms were prepared using a constant-depth film fermenter and a reconstituted human epidermis model. In vivo biofilms were also studied using biopsy samples from non-infected chronic venous leg ulcers. The specificity of peptide nucleic acid (PNA) probes for the target organisms was confirmed using mixed preparations of planktonic bacteria and multiplex PNA probing. Identification and location of individual bacterial species within multi-species biofilms demonstrated that P. aeruginosa was predominant. CLSM revealed clustering of individual species within mixed-species biofilms. FISH analysis of archive chronic wound biopsy sections showed bacterial presence and allowed bacterial load to be determined. The application of this standardized procedure makes available an assay for identification of single- or multi-species bacterial populations in tissue biopsies. The technique provides a reliable tool to study bacterial biofilm formation and offers an approach to assess targeted biofilm disruption strategies in vivo.