Search

Bacterial biofilms can cause serious health care complications associated with increased morbidity and mortality. There is an urge to discover and develop new biofilm inhibitors from natural products or by modifying natural compounds or understanding the modes of action of existing compounds. Cinnamaldehyde (CAD), one of the major components of cinnamon oil, has been demonstrated to act as an antimicrobial agent against a number of Gram-negative and Gram-positive pathogens, including Pseudomonas aeruginosa, Helicobacter pylori and Listeria monocytogenes. Despite the mechanism of action of CAD against the model organism P. aeruginosa being undefined, based on its antimicrobial properties, we hypothesized that it may disrupt preformed biofilms of P. aeruginosa. The minimum inhibitory concentration (MIC) of CAD for planktonic P. aeruginosa was determined to be 11.8 mM. Membrane depolarization assays demonstrated disruption of the transmembrane potential of P. aeruginosa. CAD at 5.9 mM (0.5 MIC) disrupted preformed biofilms by 75.6 % and 3 mM CAD (0.25 MIC) reduced the intracellular concentrations of the secondary messenger, bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP), which controls P. aeruginosa biofilm formation. The swarming motility of P. aeruginosa was also reduced by CAD in a concentration-dependent manner. Collectively, these findings show that sub-MICs of CAD can disrupt biofilms and other surface colonization phenotypes through the modulation of intracellular signalling processes.

Two new antimicrobial agents, neryl ferulate (1) and neryl p-coumarate (2), were identified using bioassay-guided isolation from the leaves of Eremophila longifolia, which is a medicinal plant used by some Australian Aboriginal communities. Although gradual autoxidation of the nerol subunit hindered the initial attempts to purify and characterize 1 and 2, it was found that the autoxidation could be stopped through storage under argon at -20 °C. Biological evaluation showed that neryl ferulate (1) had moderate activity against various Gram-positive bacteria, while neryl p-coumarate (2) was active only against Enterococcus faecium.

The broad-spectrum activity of antimicrobial peptides (AMPs) and low probability of development of host resistance make them excellent candidates as novel bio-control agents. A number of AMPs are found to be cationic, and a small proportion of these are tryptophan-rich. The puroindolines (PIN) are small, basic proteins found in wheat grains with proposed roles in biotic defence of seeds and seedlings. Synthetic peptides based on their unique tryptophan-rich domain (TRD) display antimicrobial properties. Bacterial endospores and biofilms are highly resistant cells, with significant implications in both medical and food industries. In this study, the cationic PIN TRD-based peptides PuroA (FPVTWRWWKWWKG-NH2 ) and Pina-M (FSVTWRWWKWWKG-NH2 ) and the related barley hordoindoline (HIN) based Hina (FPVTWRWWTWWKG-NH2 ) were tested for effects on planktonic cells and biofilms of the common human pathogens including Pseudomonas aeruginosa, Listeria monocytogenes and the non-pathogenic Listeria innocua. All peptides showed significant bactericidal activity. Further, PuroA and Pina-M at 2 × MIC prevented initial biomass attachment by 85-90% and inhibited >90% of 6-h preformed biofilms of all three organisms. However Hina, with a substitution of Lys-9 with uncharged Thr, particularly inhibited Listeria biofilms. The PIN based peptides were also tested against vegetative cells and endospores of Bacillus subtilis. The results provided evidence that these tryptophan-rich peptides could kill B. subtilis even in sporulated state, reducing the number of viable spores by 4 log units. The treated spores appeared withered under scanning electron microscopy. The results establish the potential of these tryptophan-rich peptides in controlling persistent pathogens of relevance to food industries and human health. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd., (Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.)

Hypothesis: Solid lipid nanoparticles (SLNs) produced by conventional microemulsion techniques using thermal heat have specific limitations (e.g. high polydispersity, instability and low encapsulation). Replacing thermal heat with microwave heat may produce SLNs which overcome some of these limitations., Experiments: Stearic acid-based SLNs prepared with Tween® 20 as the emulsifier were chosen as the optimum formulation to encapsulate and potentially deliver the antibacterial drug tetracycline. All formulations were characterized for their particle size, zeta potential, encapsulation efficiency, loading capacity, thermal and X-ray diffraction analyses. Short-term stability and in vitro drug studies were also performed., Findings: Microwave heating helps to overcome several disadvantages associated with thermal heating (nonuniform, inefficient and slow) and results in improved particle characteristics. There is thus the potential for new opportunities in the development of colloidal carriers. The particle sizes of microwave-produced SLNs were in the desired nanometer range (200-250 nm) with both lower size and lower polydispersity than the conventional SLNs. We take this as an indication of improved stability; however zeta potential measurements were not different, indicating similar stability. True stability testing (visual observation with time) did show that the microwave-induced SLNs were found to be more stable, particularly when refrigerated. The microwave-produced SLNs also demonstrated improved encapsulation efficiency and loading capacity. Thermal and diffraction analysis confirmed a lowered crystallinity of stearic acid with successful incorporation of tetracycline into the SLNs. In vitro release studies indicated that, after an initial burst release, SLNs could provide prolonged release of tetracycline. The presence of tetracycline and non-toxicity of carriers towards microbes was confirmed by antimicrobial susceptibility tests., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The plate-hole diffusion assay is an invaluable screening tool to evaluate the antibacterial potential of natural products. It relies on the diffusion of test material from pre-cut wells through agar seeded with bacteria. Samples that are capable of inhibiting bacterial growth will produce a clear zone surrounding their well. For the evaluation of antifungal activity of natural products, we describe the broth microdilution method. This assay is performed using a 96 well microtiter tray containing fungal inoculum, test medium and natural product material. Samples demonstrating antifungal activity will prevent any discernible growth as detected visually. A disk diffusion assay, utilizing the pigmented indicator strain Chromobacterium violaceum, is described here for the screening of natural products for anti quorum sensing activity. Inhibition of quorum sensing results in growth of non-pigmented bacteria.

Aim of the Study: The leaves of Planchonia careya (F. Muell.) R. Knuth (Lecythidaceae) have been traditionally implemented in the treatment of wounds by the indigenous people of northern Australia, although the compounds responsible for the medicinal properties have not been identified. In this study, antibacterial compounds from the leaf extracts were isolated and characterized, and the biological activity of each compound was assessed., Materials and Methods: Compounds were isolated from the leaf extracts using HPLC-piloted activity-guided fractionation. The minimum inhibitory concentrations (MICs) were assessed with plate-hole diffusion assays, and the cytotoxicity was determined with MTT assays using monkey kidney epithelial (MA104) cells., Results: Six known compounds were isolated from the leaf extracts and were identified as 1, (+)-gallocatechin; 2, gallocatechin-(4alpha-->8)-gallocatechin; 3, 9(S)-hydroxy-10E,12Z-octadecadienoic acid (alpha-dimorphecolic acid); 4, 2alpha,3beta,24-trihydroxyolean-12-en-28-oic acid (hyptatic acid-A); 5, 3beta-O-cis-p-coumaroyltormentic acid; and 6, 3beta-O-trans-p-coumaroyltormentic acid. Compounds 5 and 6 were weakly selective for vancomycin-resistant Enterococcus (VRE) compared with eukaryotic cells, with an MIC of 59.4microg/mL and a 50% inhibitory concentration (IC(50)) of 72.0microg/mL for MA104 cells., Conclusions: The isolation of six antibacterial compounds from the leaves of Planchonia careya validates the use of this species as a topical wound-healing remedy.

Activity-guided fractionation was used to determine the antibacterial component of an ethanolic extract of the leaves of an Australian native medicinal plant, Eremophila duttonii F. Muell. (Myoporaceae). The extract, previously shown to have activity against Gram positive bacteria, was shown to have activity against additional Gram positive bacteria, including Clostridium perfringens, C. sporogenes and Listeria monocytogenes. Thin layer chromatography (TLC) was used to separate the extract into seven coloured fractions in visible light, one of which was shown by bioautography to contain antibacterial activity. Recovery of the component from the TLC plate and testing for antibacterial activity using a plate-hole diffusion assay supported this result. The purity of the component was verified by high-performance liquid chromatography and a time-kill experiment indicated that the purified component showed identical bactericidal activity to the whole extract. TLC spray reagents indicated that the component was a sterol, terpene or sugar but not a flavonoid, while the pigmented nature suggested a carotenoid., (Copyright (c) 2004 John Wiley & Sons, Ltd.)

Ethanolic extracts of five traditional Australian medicinal plants, previously shown to display antibacterial activity against laboratory strains of the Gram positive bacteria Staphylococcus aureus and Enterococcus faecalis, were investigated for their abilities to inhibit clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE). Using plate-hole diffusion assays, the following results were obtained: (a) extract from the leaves of Eremophila alternifolia (Myoporaceae) showed activity against MRSA; (b) extract from the leaves of Acacia kempeana (Mimosaceae) showed incomplete inhibition of VRE; (c) extracts from the leaves of Amyema quandong (Loranthaceae) and Eremophila duttonii (Myoporaceae) were active against both types of bacteria; (d) extract from the stem base of Lepidosperma viscidum (Cyperaceae) was active against MRSA and exhibited incomplete inhibition of VRE. All active extracts were evaluated using time-kill assays. Most of the extracts showed bactericidal effects and reduced the number of viable cells by 4-6 logs within four hours, while the extracts from Acacia kempeana leaves and Lepidosperma viscidum stem base exhibited bacteriostatic activity against VRE. The extract from the leaves of Eremophila duttonii was the most active and reduced the number of viable cells of MRSA and VRE to undetectable levels within 1 hour.

Fifty-six ethanolic extracts of various parts of 39 plants used in traditional Australian Aboriginal medicine were investigated for their antibacterial activities against four Gram-positive (Bacillus cereus, Enterococcus faecalis, Staphylococcus aureus and Streptococcus pyogenes) and four Gram-negative (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Salmonella typhimurium) bacterial species. In a plate-hole diffusion assay, 12 extracts inhibited the growth of one or more of the bacteria, with five extracts showing broad spectrum antibacterial activity against Gram-positive bacteria. B. cereus was the most susceptible bacterium, with all 12 extracts displaying activity against this organism. Extracts from the leaves of Eremophila species (Myoporaceae) were the most active, with Eremophila duttonii exhibiting the greatest activity (against Gram-positive bacteria). The antibacterial effects of E. duttonii were further investigated by time-course growth assays which showed that significant growth inhibition was observed in cultures incubated in the presence of the extract within 1 h for B. cereus, E. faecalis and S. aureus and 2 h for S. pyogenes.