Plasma activated water pre-treatment substantially enhances phage activity against Proteus mirabilis biofilms.

The ongoing antimicrobial resistance crisis has incentivised research into alternative antibacterial and antibiofilm agents. One of them is plasma-activated water (PAW), which is produced by exposing water to a cold plasma discharge. This process generates a diverse array of reactive oxygen and nitrogen species (ROS/RNS) with antimicrobial properties. Another intensively studied class of alternative antimicrobials are bacteriophages, attracting attention due to their specificity and strong antibacterial activity. As combinations of different types of antimicrobials are known to often exhibit synergistic interactions, in this study we investigated the combined use of cold atmospheric-pressure plasma-activated water and the bacteriophage vB_PmiS_PM-CJR against Proteus mirabilis biofilms as a potential option for treatment of catheter-associated urinary tract infections (CAUTIs). We compared the effect of two cold plasma discharge setups for PAW production on its antimicrobial efficacy against P. mirabilis planktonic and biofilm cultures. Next, we assessed the stability of the phage vB_PmiS_PM-CJR in PAW. Finally, we tested the antimicrobial activity of the phages and PAW against biofilms, both individually and in combinations. Our findings demonstrate that the combination of PAW with phage is more effective against biofilms compared to individual treatments, being able to reduce the number of biofilm-embedded cells by approximately 4 log. We were also able to show that the order of treatment plays an important role in the anti-biofilm activity of the phage-PAW combination, as the exposure of the biofilm to PAW prior to phage administration results in a stronger effect than the reverse order. This research underlines PAW's ability to potentiate phage activity, showcasing a considerable reduction in biofilm viability and biomass. Additionally, it contributes to the growing body of evidence supporting the use of phage-based combinatorial treatments. Overall, this sequential treatment strategy demonstrates the potential of leveraging multiple approaches to address the mounting challenge of antibiotic resistance and offers a promising avenue for enhancing the efficacy of CAUTI management.

Climate change as a challenge for pharmaceutical storage and tackling antimicrobial resistance.

The rise of antimicrobial resistance (AMR) remains a pressing global health challenge. Infections that were once easily treatable with first-line antimicrobials are becoming increasingly difficult to manage. This shift directly threatens the wellness of humans, animals, plants, and the environment. While the AMR crisis can be attributed to a myriad of factors, including lack of infection prevention and control measures, over-prescription of antimicrobials, patient non-compliance, and the misuse of antimicrobials, one aspect that has garnered less attention is the role of storage conditions of these medicines. The way medications, particularly antimicrobials, are transported and stored until the point of use can influence their efficacy and, subsequently, may impact the development of resistant microbial strains. This review delves deeper into the often-overlooked domain of climate change (CC) and antimicrobial storage practices and the potential effects. Inappropriate storage conditions, such as exposure to extreme temperatures, humidity or light, can degrade the potency of antimicrobials. When these compromised medicines are administered to patients or animals alike, they may not effectively eradicate the targeted pathogens, leading to partial survival of the pathogens. These surviving pathogens, having been exposed to sub-lethal doses, are more likely to evolve and develop resistance mechanisms. The review discusses the mechanism underlying this and underscores the implications of antimicrobial storage practices in relation to two of the most pressing global health challenges: AMR and CC. The review also presents specific case studies and highlights the importance of monitoring storage practices and supply chain surveillance. Furthermore, the importance of deploying genomic tools to understand the potential impact of storage conditions on the development of AMR is discussed, and antimicrobial storage highlighted as a crucial part of comprehensive strategies in the fight against AMR.

Optimising intracellular delivery of antibiotic loaded PLGA nanoparticles to macrophages.

Bacteria can evade antimicrobial therapy by hiding inside host cells such as macrophages. Here we examine the ability of PLGA nanoparticles to deliver antibiotics to intracellular bacteria, specifically focusing upon the impact of nanoparticle size. Different sized Rhodamine-B conjugated PLGA nanoparticles were synthesized and uptake examined in two macrophage cell lines, as well as different epithelial cells, to determine the optimal properties for macrophage uptake. These studies demonstrate macrophages display a consistent increase in uptake with increased PLGA nanoparticle diameter. In a bacteria-macrophage co-culture model, we then examined the efficacy of different sized antibiotic-loaded PLGA nanoparticles against intracellular infections with K. pneumoniae and S. aureus. Increasing the size of antibiotic-loaded PLGA nanoparticles significantly increased their potency against intracellular K. pneumoniae. However, this was not observed for S. aureus, potentially due to the observation these nanoparticles failed to access the compartment in which S. aureus reside. This work demonstrates for the first time that increasing the size of antibiotic-loaded PLGA nanoparticles can significantly enhance antimicrobial efficacy against K. pneumoniae intracellular macrophage infections. However, our S. aureus studies indicate this is not a 'one size fits all' approach for all intracellular infections.

A novel bioinformatic method for the identification of antimicrobial peptides in metagenomes.

AIMS: This study aimed to develop a new bioinformatic approach for the identification of novel antimicrobial peptides (AMPs), which did not depend on sequence similarity to known AMPs held within databases, but on structural mimicry of another antimicrobial compound, in this case an ultrashort, synthetic, cationic lipopeptide (C12-OOWW-NH2). METHODS AND RESULTS: When applied to a collection of metagenomic datasets, our outlined bioinformatic method successfully identified several short (8-10aa) functional AMPs, the activity of which was verified via disk diffusion and minimum inhibitory concentration assays against a panel of 12 bacterial strains. Some peptides had activity comparable to, or in some cases, greater than, those from published studies that identified AMPs using more conventional methods. We also explored the effects of modifications, including extension of the peptides, observing an activity peak at 9-12aa. Additionally, the inclusion of a C-terminal amide enhanced activity in most cases. Our most promising candidate (named PB2-10aa-NH2) was thermally stable, lipid-soluble, and possessed synergistic activity with ethanol but not with a conventional antibiotic (streptomycin). CONCLUSIONS: While several bioinformatic methods exist to predict AMPs, the approach outlined here is much simpler and can be used to quickly scan huge datasets. Searching for peptide sequences bearing structural similarity to other antimicrobial compounds may present a further opportunity to identify novel AMPs with clinical relevance, and provide a meaningful contribution to the pressing global issue of AMR.

Rosin as a Natural Alternative for the effective disinfection of ESKAPE Pathogens and Clostridioides difficile spores.

AIM: Hospital-acquired infections (HAIs) caused by antimicrobial-resistant ESKAPE pathogens are a significant concern for the healthcare industry, with an estimated cost of up to ${\$}$45 billion per year in the US alone. Clostridioides difficile is an additional opportunistic pathogen that also poses a serious threat to immunocompromised patients in hospitals. Infections caused by these pathogens lead to increased hospital stays and repeated readmission, resulting in a significant economic burden. Disinfectants and sporicidals are essential to reduce the risk of these pathogens in hospitals, but commercially available products can have a number of disadvantages including inefficacy, long contact times, short shelf lives, and operator health hazards. In this study we evaluated the effectiveness of Rosin (a natural substance secreted by coniferous trees as a defence mechanism against wounds in tree bark) and its commercial derivative Rosetax-21 as disinfectants and sporicidal against the six ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) and spore preparations from Clostridioides difficile. METHODS AND RESULTS: Both Rosin and Rosetax-21 were tested under simulated clean and dirty conditions (with BSA) against the ESKAPE pathogens, and C. difficile spore preparations. In clean conditions, Rosin (5% weight/volume: w/v) demonstrated significant efficacy against five of the ESKAPE pathogens, with A. baumannii and E. faecium being the most susceptible, and K. pneumoniae the most resistant, showing only a one-log reduction after a 5 min treatment. However, in dirty conditions, all pathogens including K. pneumoniae exhibited at least a 3-log reduction to Rosin within 5 min. Rosetax-21 (5% w/v) was found to be less effective than Rosin in clean conditions, a trend that was exacerbated in the presence of BSA. Additionally, both Rosin and Rosetax-21 at 2.5% (w/v) achieved complete eradication of C. difficile spores when combined with 0.5% glutaraldehyde, though their standalone sporicidal activity was limited. CONCLUSIONS: The findings from this study highlight the potential of Rosin and Rosetax-21 as both bactericidal and sporicidal disinfectants, with their efficacy varying based on the conditions and the pathogens tested. This presents an avenue for the development of novel healthcare disinfection strategies, especially against HAIs caused by antimicrobial-resistant ESKAPE pathogens and C. difficile.

Curcumin nanocrystals-in-nanofibres as a promising platform for the management of periodontal disease.

It is estimated that nearly a half of the world's population over 30 years old suffer from some kind of periodontal disease (PD). Although preventable, PD can pose a significant health burden to patients, causing from pain and discomfort to disfigurement and death. The management of PD often requires surgical procedures accompanied of systemic antibiotic and anti-inflammatory treatments. Curcumin (CUR), a potent anti-inflammatory and antimicrobial active, has shown great promise in the management of PD; however, its effects are often limited by its low bioavailability. In this work, we report the development of electrospun nanofibres (NFs) loaded with CUR nanocrystals (NCs) for the management of PD. NCs of 100 nm were obtained by media milling and loaded into dissolving polyvinyl alcohol NFs using electrospinning. The resultant NCs-in-NFs dissolved in water spontaneously, releasing NCs with a particle size of âˆ¼120 nm. The physiochemical characterisation of the systems indicated the absence of chemical interactions between drug and polymer, and nanofibres with an amorphous nature. In vitro release profiles demonstrated that the NCs had a significantly higher dissolution rate (∼100 % at day 40) than the control group (approximately 6 % at day 40), which consisted of NFs containing a physical mixture of the drug and stabiliser. Finally, mucosal deposition studies demonstrated a 10-fold higher capacity of the novel NCs-in-NFs system to deposit CUR ex vivo using excised neonatal porcine mucosal tissue, when compared to the control group.

Assessment of Cold Atmospheric Pressure Plasma (CAPP) Treatment for Degradation of Antibiotic Residues in Water.

The presence of antibiotic residues in water is linked to the emergence of antibiotic resistance globally and necessitates novel decontamination strategies to minimize antibiotic residue exposure in both the environment and food. A holistic assessment of cold atmospheric pressure plasma technology (CAPP) for ß-lactam antibiotic residue removal is described in this study. CAPP operating parameters including plasma jet voltage, gas composition and treatment time were optimized, with highest ß-lactam degradation efficiencies obtained for a helium jet operated at 6 kV. Main by-products detected indicate pH-driven peroxidation as a main mechanism of CAPP-induced decomposition of ß-lactams. No in vitro hepatocytotoxicity was observed in HepG2 cells following exposure to treated samples, and E. coli exposed to CAPP-degraded ß-lactams did not exhibit resistance development. In surface water, over 50% decrease in antibiotic levels was achieved after only 5 min of treatment. However, high dependence of treatment efficiency on residue concentration, pH and presence of polar macromolecules was observed.

Quorum Sensing in Halorubrum saccharovorum Facilitates Cross-Domain Signaling between Archaea and Bacteria.

Quorum Sensing (QS) is a well-studied intercellular communication mechanism in bacteria, regulating collective behaviors such as biofilm formation, virulence, and antibiotic resistance. However, cell-cell signaling in haloarchaea remains largely unexplored. The coexistence of bacteria and archaea in various environments, coupled with the known cell-cell signaling mechanisms in both prokaryotic and eukaryotic microorganisms and the presence of cell-cell signaling mechanisms in both prokaryotic and eukaryotic microorganisms, suggests a possibility for haloarchaea to possess analogous cell-cell signaling or QS systems. Recently, N-acylhomoserine lactone (AHL)-like compounds were identified in haloarchaea; yet, their precise role-for example, persister cell formation-remains ambiguous. This study investigated the capacity of crude supernatant extract from the haloarchaeon Halorubrum saccharovorum CSM52 to stimulate bacterial AHL-dependent QS phenotypes using bioreporter strains. Our findings reveal that these crude extracts induced several AHL-dependent bioreporters and modulated pyocyanin and pyoverdine production in Pseudomonas aeruginosa. Importantly, our study suggests cross-domain communication between archaea and bacterial pathogens, providing evidence for archaea potentially influencing bacterial virulence. Using Thin Layer Chromatography overlay assays, lactonolysis, and colorimetric quantification, the bioactive compound was inferred to be a chemically modified AHL-like compound or a diketopiperazine-like molecule, potentially involved in biofilm formation in H. saccharovorum CSM52. This study offers new insights into putative QS mechanisms in haloarchaea and their potential role in interspecies communication and coordination, thereby enriching our understanding of microbial interactions in diverse environments.

Cold atmospheric pressure plasma-antibiotic synergy in Pseudomonas aeruginosa biofilms is mediated via oxidative stress response.

Cold atmospheric-pressure plasma (CAP) has emerged as a potential alternative or adjuvant to conventional antibiotics for the treatment of bacterial infections, including those caused by antibiotic-resistant pathogens. The potential of sub-lethal CAP exposures to synergise conventional antimicrobials for the eradication of Pseudomonas aeruginosa biofilms is investigated in this study. The efficacy of antimicrobials following or in the absence of sub-lethal CAP pre-treatment in P. aeruginosa biofilms was assessed. CAP pre-treatment resulted in an increase in both planktonic and biofilm antimicrobial sensitivity for all three strains tested (PAO1, PA14, and PA10548), with both minimum inhibitory concentrations (MICs) and minimum biofilm eradication concentrations (MBECs) of individual antimicrobials, being significantly reduced following CAP pre-treatment of the biofilm (512-fold reduction with ciprofloxacin/gentamicin; and a 256-fold reduction with tobramycin). At all concentrations of antimicrobial used, the combination of sub-lethal CAP exposure and antimicrobials was effective at increasing time-to-peak metabolism, as measured by isothermal microcalorimetry, again indicating enhanced susceptibility. CAP is known to damage bacterial cell membranes and DNA by causing oxidative stress through the in situ generation of reactive oxygen and nitrogen species (RONS). While the exact mechanism is not clear, oxidative stress on outer membrane proteins is thought to damage/perturb cell membranes, confirmed by ATP and LDH leakage, allowing antimicrobials to penetrate the bacterial cell more effectively, thus increasing bacterial susceptibility. Transcriptomic analysis, reveals that cold-plasma mediated oxidative stress caused upregulation of P. aeruginosa superoxide dismutase, cbb3 oxidases, catalases, and peroxidases, and upregulation in denitrification genes, suggesting that P. aeruginosa uses these enzymes to degrade RONS and mitigate the effects of cold plasma mediated oxidative stress. CAP treatment also led to an increased production of the signalling molecule ppGpp in P. aeruginosa, indicative of a stringent response being established. Although we did not directly measure persister cell formation, this stringent response may potentially be associated with the formation of persister cells in biofilm cultures. The production of ppGpp and polyphosphate may be associated with protein synthesis inhibition and increase efflux pump activity, factors which can result in antimicrobial tolerance. The transcriptomic analysis also showed that by 6 h post-treatment, there was downregulation in ribosome modulation factor, which is involved in the formation of persister cells, suggesting that the cells had begun to resuscitate/recover. In addition, CAP treatment at 4 h post-exposure caused downregulation of the virulence factors pyoverdine and pyocyanin; by 6 h post-exposure, virulence factor production was increasing. Transcriptomic analysis provides valuable insights into the mechanisms by which P. aeruginosa biofilms exhibits enhanced susceptibility to antimicrobials. Overall, these findings suggest, for the first time, that short CAP sub-lethal pre-treatment can be an effective strategy for enhancing the susceptibility of P. aeruginosa biofilms to antimicrobials and provides important mechanistic insights into cold plasma-antimicrobial synergy. Transcriptomic analysis of the response to, and recovery from, sub-lethal cold plasma exposures in P. aeruginosa biofilms improves our current understanding of cold plasma biofilm interactions.

Exploring halophilic environments as a source of new antibiotics.

Microbial natural products from microbes in extreme environments, including haloarchaea, and halophilic bacteria, possess a huge capacity to produce novel antibiotics. Additionally, enhanced isolation techniques and improved tools for genomic mining have expanded the efficiencies in the antibiotic discovery process. This review article provides a detailed overview of known antimicrobial compounds produced by halophiles from all three domains of life. We summarize that while halophilic bacteria, in particular actinomycetes, contribute the vast majority of these compounds the importance of understudied halophiles from other domains of life requires additional consideration. Finally, we conclude by discussing upcoming technologies- enhanced isolation and metagenomic screening, as tools that will be required to overcome the barriers to antimicrobial drug discovery. This review highlights the potential of these microbes from extreme environments, and their importance to the wider scientific community, with the hope of provoking discussion and collaborations within halophile biodiscovery. Importantly, we emphasize the importance of bioprospecting from communities of lesser-studied halophilic and halotolerant microorganisms as sources of novel therapeutically relevant chemical diversity to combat the high rediscovery rates. The complexity of halophiles will necessitate a multitude of scientific disciplines to unravel their potential and therefore this review reflects these research communities.

Microbial communities of halite deposits and other hypersaline environments.

Large regions of Earth's surface are underlain by salt deposits that evaporated from ancient oceans and are populated by extreme halophilic microbes. While the microbiology of ancient evaporites has been well studied, the ecology of halite deposits and more recently formed NaCl "salticle" stalactite structures (speleothems) in a Triassic halite mine are less well characterized. The microbiome of Kilroot Salt Mine was profiled using conventional and enhanced culturing techniques. From this, 89 halophilic archaeal isolates from six known genera, and 55 halophilic or halotolerant bacterial isolates from 18 genera were obtained. Culture-independent metagenomic approaches also revealed that culturing techniques were inadvertently biased toward specific taxa, and the need for optimized isolation procedures are required to enhance cultivation diversity. Speleothems formed from saturated brines are unique structures that have the potential to entomb haloarchaea cells for thousands of years within fluid inclusions. The presence of such fluid inclusions, alongside the high abundance of genes related to glycerol metabolism, biofilm formation, and persister cell formation is highly suggestive of an environmental niche that could promote longevity and survivability. Finally, previous studies reporting the discovery of novel biocatalysts from the Kilroot mine microbiome, suggests that this environment may be an untapped source of chemical diversity with high biodiscovery potential.

Extracts of Sida cordifolia contain polysaccharides possessing immunomodulatory activity and rosmarinic acid compounds with antibacterial activity.

BACKGROUND: The overuse of antibiotics has led to increased antimicrobial resistance, but plant-derived biological response modifiers represent a potential alternative to these drugs. This investigation examined the immunomodulatory and antibacterial activities of Sida cordifolia (used in ethnomedicinal systems to treat infectious disease). METHODS: Successive extractions were performed from the roots of these plants in hexane, chloroform, methanol and water. Immunomodulatory activity was determined in a series of experiments measuring the responses of splenocytes, macrophages and an in vivo model of innate immunity (Galleria mellonella). Antibacterial activity was assessed by determining minimum inhibitory/bactericidal concentrations (MIC/MBCs) for various Gram-positive and Gram-negative bacterial strains. RESULTS: Immunomodulatory activity was confined to the aqueous extract, and further fractionation and biochemical analysis yielded a highly potent polysaccharide-enriched fraction (SCAF5). SCAF5 is a complex mixture of different polysaccharides with multiple immunomodulatory effects including immune cell proliferation, antibody secretion, phagocytosis, nitric oxide production, and increased expression of pro-inflammatory cytokines. Furthermore, Galleria mellonella pre-treated with SCAF5 produced more haemocytes and were more resistant (P < 0.001) to infection with methicillin-resistant Staphylococcus aureus (MRSA) with a 98% reduction in bacterial load in pre-treated larvae compared to the negative control. The antibacterial activity of Sida cordifolia was confined to the methanolic fraction. Extensive fractionation identified two compounds, rosmarinic acid and its 4-O-ß-d-glucoside derivative, which had potent activity against Gram-positive antibiotic-resistant bacteria, including MRSA. CONCLUSIONS: Sida cordifolia counters bacterial infections through a dual mechanism, and immunomodulatory polysaccharides from this plant should be isolated and characterised to realise their potential as anti-infective agents. Such properties could be developed as an antibiotic alternative (1) in the clinic and (2) alternative growth promoter for the agri-food industry.

Draft Genome Sequences of Halobacterium sp. Strains KA-4 and KA-6, Two Extremely Halophilic Archaea Isolated from a Triassic Salt Deposit in Northern Ireland.

Here, we report the draft genome sequences of Halobacterium sp. strains KA-4 and KA-6. These extremely halophilic archaea were isolated from a Triassic halite deposit in Northern Ireland. Based on 16S sequence identity, they were deemed to be closely related strains of Halobacterium noricense but with some notable phenotypic differences.

Novel Phage-Derived Depolymerase with Activity against Proteus mirabilis Biofilms.

The adherence of Proteus mirabilis to the surface of urinary catheters leads to colonization and eventual blockage of the catheter lumen by unique crystalline biofilms produced by these opportunistic pathogens, making P. mirabilis one of the leading causes of catheter-associated urinary tract infections. The Proteus biofilms reduce efficiency of antibiotic-based treatment, which in turn increases the risk of antibiotic resistance development. Bacteriophages and their enzymes have recently become investigated as alternative treatment options. In this study, a novel Proteus bacteriophage (vB_PmiS_PM-CJR) was isolated from an environmental sample and fully characterized. The phage displayed depolymerase activity and the subsequent genome analysis revealed the presence of a pectate lyase domain in its tail spike protein. The protein was heterologously expressed and purified; the ability of the purified tail spike to degrade Proteus biofilms was tested. We showed that the application of the tail spike protein was able to reduce the adherence of bacterial biofilm to plastic pegs in a MBEC (minimum biofilm eradication concentration) assay and improve the survival of Galleria mellonella larvae infected with Proteus mirabilis. Our study is the first to successfully isolate and characterize a biofilm depolymerase from a Proteus phage, demonstrating the potential of this group of enzymes in treatment of Proteus infections.

Antibiotic Therapy and the Gut Microbiome: Investigating the Effect of Delivery Route on Gut Pathogens.

The contribution of the gut microbiome to human health has long been established, with normal gut microbiota conferring protection against invasive pathogens. Antibiotics can disrupt the microbial balance of the gut, resulting in disease and the development of antimicrobial resistance. The effect of antibiotic administration route on gut dysbiosis remains under-studied to date, with conflicting evidence on the differential effects of oral and parenteral delivery. We have profiled the rat gut microbiome following treatment with commonly prescribed antibiotics (amoxicillin and levofloxacin), via either oral or intravenous administration. Fecal pellets were collected over a 13-day period and bacterial populations were analyzed by 16S rRNA gene sequencing. Significant dysbiosis was observed in all treatment groups, regardless of administration route. More profound dysbiotic effects were observed following amoxicillin treatment than those with levofloxacin, with population richness and diversity significantly reduced, regardless of delivery route. The effect on specific taxonomic groups was assessed, revealing significant disruption following treatment with both antibiotics. Enrichment of a number of groups containing known gut pathogens was observed, in particular, with amoxicillin, such as the family Enterobacteriaceae. Depletion of other commensal groups was also observed. The degree of dysbiosis was significantly reduced toward the end of the sampling period, as bacterial populations began to return to pretreatment composition. Richness and diversity levels appeared to return to pretreatment levels more quickly in intravenous groups, suggesting convenient parenteral delivery systems may have a role to play in reducing longer term gut dysbiosis in the treatment of infection.

Microbiology of a NaCl stalactite 'salticle' in Triassic halite.

Large regions of Earth's surface are underlain by salt deposits that evaporated from ancient oceans and are populated by extreme halophilic microbes. Some of these halophiles may have been preserved over geological timescales within hypersaline fluid inclusions, but ingresses of water and/or anthropogenic activities can lead to the formation of alternative habitats, including NaCl stalactites or other speleothems. While the microbiology of ancient evaporites has been well studied, the ecology of these recently formed structures is less-well understood. Here, the microbiology of a NaCl stalactite ('salticle') in a Triassic halite mine is characterized. The specific aims were to determine the presence of fluid inclusions, determine the microbial structure of the salticle compared with a nearby brine-pool and surficial soil, and characterize the ecophysiological capabilities of this unique ecosystem. The salticle contained fluid inclusions, and their microbiome was composed of Euryarchaetota, Proteobacteria, and Actinobacteria, with Haloarchaea in greater abundance than brine-pool or soil microbiomes. The salticle metagenome exhibited a greater abundance of genes involved in osmoregulation, anaerobic respiration, UV resistance, oxidative stress, and stress-protein synthesis relative to the soil microbiome. We discuss the potential astrobiological implications of salticles as enclosed salt-saturated habitats that are protected from ionizing radiation and have a stable water activity.

Characterization of ionic liquid cytotoxicity mechanisms in human keratinocytes compared with conventional biocides.

The ability to chemically modify ionic liquids (ILs) has led to an expansion in interest in their use in a diversity of applications, not least as antimicrobials and biocides. Relatively little is known about cytotoxicity mechanisms of ILs in comparison to other biocides currently in widespread use, as well as their practical significance for the ecological environment and human health. Using NCTC 2544 and HaCat human keratinocyte cells, this study aimed to characterize cytotoxicity rates and mechanisms of a range of ILs. Using both lactate dehydrogenase (LDH) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) based cytotoxicity assays, it was confirmed that at biocide-relevant concentrations, ILs with longer alkyl chains exhibited greater biocidal activity than those with shorter alkyl chains, with comparable activity to the commonly used biocides chlorhexidine, benzalkonium chloride and cetylpyridinium chloride, at relevant in-use biocide concentrations. Mode of cell death, measured using fluorescence-activated cell sorting (FACS) and caspase 3/7 activity, determined necrosis to be the primary cytotoxic mechanism at higher concentrations of the biocides stated above, and with ILs [C14MIM]Cl and [C14quin]Br, with apoptosis observed at borderline necrotic concentrations. Perhaps most interestingly, modification of anion had a significant effect on cytotoxicity. The use of N[SO2CF3] as an anion to [C16MIM] attenuated cytotoxicity 10-fold in comparison to other anions, suggesting cytotoxicity may also be a tuneable property when using ILs as biocides.

Antimicrobial 3D Printed Objects in the Fight Against Pandemics.

The uncharted nature of the COVID-19 pandemic has caused uncertainty globally, resulting in many health care professionals and key-workers being left with supply shortages in medical consumables and personal protective equipment, exacerbated by supply line issues and in some cases delays resulting from governmental policies. 3D printing (3DP) has played an important role in providing essential items to hospitals and the wider communities, such as visors, face masks, and ventilator components. This short-review article covers the potential of antimicrobial materials in the manufacturing of 3DP essential products, as an approach for added protection against pandemics.

Biomolecules as Model Indicators of In Vitro and In Vivo Cold Plasma Safety.

The potential applications for cold plasma in medicine are extensive, from microbial inactivation and induction of apoptosis in cancer cells to stimulating wound healing and enhancing the blood coagulation cascade. The safe bio-medical application of cold plasma and subsequent effect on complex biological pathways requires precision and a distinct understanding of how physiological redox chemistry is manipulated. Chemical modification of biomolecules such as carbohydrates, proteins, and lipids treated with cold plasma have been characterized, however, the context of how alterations of these molecules affect cell behavior or in vivo functionality has not been determined. Thus, this study examines the cytotoxic and mutagenic effects of plasma-treated molecules in vitro using CHO-K1 cells and in vivo in Galleria mellonella larvae. Specifically, albumin, glucose, cholesterol, and arachidonic acid were chosen as representative biomolecules, with established involvement in diverse bioprocesses including; cellular respiration, intracellular transport, cell signaling or membrane structure. Long- and short-term effects depended strongly on the molecule type and the treatment milieu indicating the impact of chemical and physical modifications on downstream biological pathways. Importantly, absence of short-term toxicity did not always correlate with absence of longer-term effects, indicating the need to comprehensively assess ongoing effects for diverse biological applications.