Multifunctional host-defense peptides isolated from skin secretions of the banana tree dwelling frog Boana platanera (Hylidae; Hylinae).

Five host-defense peptides (figainin 2PL, hylin PL, raniseptin PL, plasticin PL, and peptide YL) were isolated from norepinephrine-stimulated skin secretions of the banana tree dwelling frog Boana platanera (Hylidae; Hylinae) collected in Trinidad. Raniseptin PL (GVFDTVKKIGKAVGKFALGVAKNYLNS.NH2) and figainin 2PL (FLGTVLKLGKAIAKTVVPMLTNAMQPKQ. NH2) showed potent and rapid bactericidal activity against a range of clinically relevant Gram-positive and Gram-negative ESKAPE + pathogens and Clostridioides difficile. The peptides also showed potent cytotoxic activity (LC50 values < 30 µM) against A549, MDA-MB-231 and HT29 human tumor-derived cell lines but appreciably lower hemolytic activity against mouse erythrocytes (LC50 = 262 ± 14 µM for raniseptin PL and 157 ± 16 µM for figainin 2PL). Hylin PL (FLGLIPALAGAIGNLIK.NH2) showed relatively weak activity against microorganisms but was more hemolytic. The glycine-leucine-rich peptide with structural similarity to the plasticins (GLLSTVGGLVGGLLNNLGL.NH2) and the non-cytotoxic peptide YL (YVPGVIESLL.NH2) lacked antimicrobial and cytotoxic activities. Hylin PL, raniseptinPL and peptide YL stimulated the rate of release of insulin from BRIN-BD11 clonal ß-cells at concentrations ≥100 nM. Peptide YL was the most effective (2.3-fold increase compared with basal rate at 1 µM concentration) and may represent a template for the design of a new class of incretin-based anti-diabetic drugs.

Microbial Biosurfactants: Antimicrobial Activity and Potential Biomedical and Therapeutic Exploits.

The rapid emergence of multidrug-resistant pathogens worldwide has raised concerns regarding the effectiveness of conventional antibiotics. This can be observed in ESKAPE pathogens, among others, whose multiple resistance mechanisms have led to a reduction in effective treatment options. Innovative strategies aimed at mitigating the incidence of antibiotic-resistant pathogens encompass the potential use of biosurfactants. These surface-active agents comprise a group of unique amphiphilic molecules of microbial origin that are capable of interacting with the lipidic components of microorganisms. Biosurfactant interactions with different surfaces can affect their hydrophobic properties and as a result, their ability to alter microorganisms' adhesion abilities and consequent biofilm formation. Unlike synthetic surfactants, biosurfactants present low toxicity and high biodegradability and remain stable under temperature and pH extremes, making them potentially suitable for targeted use in medical and pharmaceutical applications. This review discusses the development of biosurfactants in biomedical and therapeutic uses as antimicrobial and antibiofilm agents, in addition to considering the potential synergistic effect of biosurfactants in combination with antibiotics. Furthermore, the anti-cancer and anti-viral potential of biosurfactants in relation to COVID-19 is also discussed.

Identification of an Antimicrobial Peptide from the Venom of the Trinidad Thick-Tailed Scorpion Tityus trinitatis with Potent Activity against ESKAPE Pathogens and Clostridioides difficile.

Envenomation by the Trinidad thick-tailed scorpion Tityus trinitatis may result in fatal myocarditis and there is a high incidence of acute pancreatitis among survivors. Peptidomic analysis (reversed-phase HPLC followed by MALDI-TOF mass spectrometry and automated Edman degradation) of T. trinitatis venom led to the isolation and characterization of three peptides with antimicrobial activity. Their primary structures were established asTtAP-1 (FLGSLFSIGSKLLPGVFKLFSRKKQ.NH2), TtAP-2 (IFGMIPGLIGGLISAFK.NH2) and TtAP-3 (FFSLIPSLIGGLVSAIK.NH2). In addition, potassium channel and sodium channel toxins, present in the venom in high abundance, were identified by CID-MS/MS sequence analysis. TtAP-1 was the most potent against a range of clinically relevant Gram-positive and Gram-negative aerobes and against the anaerobe Clostridioides difficile (MIC = 3.1-12.5 µg/mL). At a concentration of 1× MIC, TtAP-1 produced rapid cell death (<15 min against Acinetobacter baumannii and Staphylococcus aureus). The therapeutic potential of TtAP-1 as an anti-infective agent is limited by its high hemolytic activity (LC50 = 18 µg/mL against mouse erythrocytes) but the peptide constitutes a template for the design of analogs that maintain the high bactericidal activity against ESKAPE pathogens but are less toxic to human cells. It is suggested that the antimicrobial peptides in the scorpion venom facilitate the action of the neurotoxins by increasing the membrane permeability of cells from either prey or predator.

Embedding Scientific Communication and Digital Capabilities in the Undergraduate Biomedical Science Curriculum.

Introduction: Scientific communication, particularly the dissemination of research findings to both the scientific community and the general public, are skills required of graduates embarking on post-graduate studies and employment within the biomedical sciences sector. The aims of this action research project were to i) co-design an online scientific communication and digital capabilities resource, constructively aligned to the learning objectives of a final year undergraduate investigative research project; ii) ensure resource flexibility for future adaptation by others iii) embed authentic scientific communication learning assessments, namely, the preparation of a lay summary and visual abstract and iv) promote students' awareness of developed digital capabilities and transferable skills through written reflection. Materials and Methods: Student engagement, self-efficacy, experiences and performance and staff perceptions (n = 15) were evaluated by a mixed methods approach. Qualitative data was gathered from focus sessions, free text responses within questionnaires and content analysis of students' written reflections (n = 104). Quantitative data from 5-point Likert responses within student questionnaires (n = 31) and analysis of student scientific and lay writing (n = 146) using the readability parameters Flesch-Kincaid Grade Level and Flesch Reading Ease were analysed using non-parametric statistical methods. Results: A learning resource was co-designed with students, staff, local, national and international contributors and valued by both students and staff, enabling students to prepare scientific communication outputs of a professional standard by application of digital, analytical and scientific communication skills. Students prepared lay summaries which were statistically (p < 0.0001) more readable than their paired scientific abstracts. Significant correlations between easier readability of lay summaries and awarded marks for the written elements of the module were noted. Students reported their digital and communication capabilities increased significantly (p < 0.0001) throughout, from limited to good/excellent and reflected on the numerous transferable skills developed during preparation of assessments, with 75% reflecting on their digital capabilities. Discussion: Undergraduate students developed, appreciated and used varied scientific communication and digital skills to articulate research findings. The embedding of such activities throughout all levels of higher education will enable students to develop their digital and scientific skills and reflect on the development of such transferable skills for application in their future careers.

Furanone loaded aerogels are effective antibiofilm therapeutics in a model of chronic Pseudomonas aeruginosa wound infection.

Almost 80% of chronic wounds have a bacterial biofilm present. These wound biofilms are caused by a range of organisms and are often polymicrobial. Pseudomonas aeruginosa is one of the most common causative organisms in wound infections and readily forms biofilms in wounds. To coordinate this, P. aeruginosa uses a process known as quorum sensing. Structural homologues of the quorum sensing signalling molecules have been used to disrupt this communication and prevent biofilm formation by Pseudomonas. However, these compounds have not yet reached clinical use. Here, we report the production and characterisation of a lyophilised PVA aerogel for use in delivering furanones to wound biofilms. PVA aerogels successfully release a model antimicrobial and two naturally occurring furanones in an aqueous environment. Furanone loaded aerogels inhibited biofilm formation in P. aeruginosa by up to 98.80%. Further, furanone loaded aerogels successfully reduced total biomass of preformed biofilms. Treatment with a sotolon loaded aerogel yielded a 5.16 log reduction in viable biofilm bound cells in a novel model of chronic wound biofilm, equivalent to the current wound therapy Aquacel AG. These results highlight the potential utility of aerogels in drug delivery to infected wounds and supports the use of biofilm inhibitory compounds as wound therapeutics.

Biological and synthetic surfactant exposure increases antimicrobial gene occurrence in a freshwater mixed microbial biofilm environment.

Aquatic habitats are particularly susceptible to chemical pollution, such as antimicrobials, from domestic, agricultural, and industrial sources. This has led to the rapid increase of antimicrobial resistance (AMR) gene prevalence. Alternate approaches to counteract pathogenic bacteria are in development including synthetic and biological surfactants such as sodium dodecyl sulfate (SDS) and rhamnolipids. In the aquatic environment, these surfactants may be present as pollutants with the potential to affect biofilm formation and AMR gene occurrence. We tested the effects of rhamnolipid and SDS on aquatic biofilms in a freshwater stream in Northern Ireland. We grew biofilms on contaminant exposure substrates deployed within the stream over 4 weeks. We then extracted DNA and carried out shotgun sequencing using a MinION portable sequencer to determine microbial community composition, with 16S rRNA analyses (64,678 classifiable reads identified), and AMR gene occurrence (81 instances of AMR genes over 9 AMR gene classes) through a metagenomic analysis. There were no significant changes in community composition within all systems; however, biofilm exposed to rhamnolipid had a greater number of unique taxa as compared to SDS treatments and controls. AMR gene prevalence was higher in surfactant-treated biofilms, although not significant, with biofilm exposed to rhamnolipids having the highest presence of AMR genes and classes compared to the control or SDS treatments. Our results suggest that the presence of rhamnolipid encourages an increase in the prevalence of AMR genes in biofilms produced in mixed-use water bodies.

Efficacy of single pass UVC air treatment for the inactivation of coronavirus, MS2 coliphage and Staphylococcus aureus bioaerosols.

There is strong evidence that SARS-CoV-2 is spread predominantly by airborne transmission, with high viral loads released into the air as respiratory droplets and aerosols from the infected subject. The spread and persistence of SARS-CoV-2 in diverse indoor environments reinforces the urgent need to supplement distancing and PPE based approaches with effective engineering measures for microbial decontamination - thereby addressing the significant risk posed by aerosols. We hypothesized that a portable, single-pass UVC air treatment device (air flow 1254 L/min) could effectively inactivate bioaerosols containing bacterial and viral indicator organisms, and coronavirus without reliance on filtration technology, at reasonable scale. Robust experiments demonstrated UVC dose dependent inactivation of Staphylococcus aureus (UV rate constant (k) = 0.098 m2/J) and bacteriophage MS2, with up to 6-log MS2 reduction achieved in a single pass through the system (k = 0.119 m2/J). The inclusion of a PTFE diffuse reflector increased the effective UVC dose by up to 34% in comparison to a standard Al foil reflector (with identical lamp output), resulting in significant additional pathogen inactivation (1-log S. aureus and MS2, p < 0.001). Complete inactivation of bovine coronavirus bioaerosols was demonstrated through tissue culture infectivity (2.4-log reduction) and RT-qPCR analysis - confirming single pass UVC treatment to effectively deactivate coronavirus to the limit of detection of the culture-based method. Scenario-based modelling was used to investigate the reduction in risk of airborne person-to-person transmission based upon a single infected subject within the small room. Use of the system providing 5 air changes per hour was shown to significantly reduce airborne viral load and maintain low numbers of RNA copies when the infected subject remained in the room, reducing the risk of airborne pathogen transmission to other room users. We conclude that the application of single-pass UVC systems (without reliance on HEPA filtration) could play a critical role in reducing the risk of airborne pathogen transfer, including SARS-CoV2, in locations where adequate fresh air ventilation cannot be implemented.

Ex Vivo Fecal Fermentation of Human Ileal Fluid Collected After Wild Strawberry Consumption Modulates Human Microbiome Community Structure and Metabolic Output and Protects Against DNA Damage in Colonic Epithelial Cells.

SCOPE: Wild strawberries (Fragaria vesca) are richer in (poly)phenols than common commercial strawberry varieties, e.g., Fragaria × ananassa. (Poly)phenols and their microbiota-derived metabolites are hypothesized to exert bioactivity within the human gut mucosa. To address this, the effects of wild strawberries are investigated with respect to their bioactivity and microbiota-modulating capacity using both in vitro and ex vivo approaches. METHODS AND RESULTS: Ileal fluids collected pre- (0h) and post-consumption (8h) of 225 g wild strawberries by ileostomates (n = 5) and also in vitro digested strawberry varieties (Fragaria vesca and Fragaria × ananassa Duchesne) supernatants are collected. Subsequent fermentation of these supernatants using an in vitro batch culture proximal colon model reveals significant treatment-specific changes in microbiome community structure in terms of alpha but not beta diversity at 24 h. Nutri-kinetic analysis reveals a significant increase in the concentration of gut microbiota catabolites, including 3-(4hydroxyphenyl)propionic acid, 3-(3-hydroxyphenyl)propanoic acid, and benzoic acid. Furthermore, post-berry ileal fermentates (24 h) significantly (p < 0.01) decrease DNA damage (% Tail DNA, COMET assay) in both HT29 cells (∼45%) and CCD 841 CoN cells (∼25%) compared to untreated controls. CONCLUSIONS: Post berry consumption fermentates exhibit increased overall levels of (poly)phenolic metabolites, which retains their bioactivity, reducing DNA damage in colonocytes.

Low-Molecular-Weight Seaweed-Derived Polysaccharides Lead to Increased Faecal Bulk but Do Not Alter Human Gut Health Markers.

Seaweeds are potentially sustainable crops and are receiving significant interest because of their rich bioactive compound content; including fatty acids, polyphenols, carotenoids, and complex polysaccharides. However, there is little information on the in vivo effects on gut health of the polysaccharides and their low-molecular-weight derivatives. Herein, we describe the first investigation into the prebiotic potential of low-molecular-weight polysaccharides (LMWPs) derived from alginate and agar in order to validate their in vivo efficacy. We conducted a randomized; placebo-controlled trial testing the impact of alginate and agar LWMPs on faecal weight and other markers of gut health and on composition of gut microbiota. We show that these LMWPs led to significantly increased faecal bulk (20-30%). Analysis of gut microbiome composition by sequencing indicated no significant changes attributable to treatment at the phylum and family level, although FISH analysis showed an increase in Faecalibacterium prausnitzii in subjects consuming agar LMWP. Sequence analysis of gut bacteria corroborated with the FISH data, indicating that alginate and agar LWMPs do not alter human gut microbiome health markers. Crucially, our findings suggest an urgent need for robust and rigorous human in vivo testing-in particular, using refined seaweed extracts.

Biofilm regulation in Clostridioides difficile: Novel systems linked to hypervirulence.

Clostridiodes difficile (C. difficile) was ranked an "urgent threat" by the Centers for Disease Control and Prevention (CDC) in 2019. C. difficile infection (CDI) is the most common healthcare-associated infection (HAI) in the United States of America as well as the leading cause of antibiotic-associated gastrointestinal disease. C. difficile is a gram-positive, rod-shaped, spore-forming, anaerobic bacterium that causes infection of the epithelial lining of the gut. CDI occurs most commonly after disruption of the human gut microflora following the prolonged use of broad-spectrum antibiotics. However, the recurrent nature of this disease has led to the hypothesis that biofilm formation may play a role in its pathogenesis. Biofilms are sessile communities of bacteria protected from extracellular stresses by a matrix of self-produced proteins, polysaccharides, and extracellular DNA. Biofilm regulation in C. difficile is still incompletely understood, and its role in disease recurrence has yet to be fully elucidated. However, many factors have been found to influence biofilm formation in C. difficile, including motility, adhesion, and hydrophobicity of the bacterial cells. Small changes in one of these systems can greatly influence biofilm formation. Therefore, the biofilm regulatory system would need to coordinate all these systems to create optimal biofilm-forming physiology under appropriate environmental conditions. The coordination of these systems is complex and multifactorial, and any analysis must take into consideration the influences of the stress response, quorum sensing (QS), and gene regulation by second messenger molecule cyclic diguanosine monophosphate (c-di-GMP). However, the differences in biofilm-forming ability between C. difficile strains such as 630 and the "hypervirulent" strain, R20291, make it difficult to assign a "one size fits all" mechanism to biofilm regulation in C. difficile. This review seeks to consolidate published data regarding the regulation of C. difficile biofilms in order to identify gaps in knowledge and propose directions for future study.

Does each bead count? A reduced-cost approach for recovering waterborne protozoa from challenge water using immunomagnetic separation.

Giardia duodenalis and Cryptosporidium spp. are two of the most prominent aetiological agents of waterborne diseases. Therefore, efficient and affordable methodologies for identifying and quantifying these parasites in water are increasingly necessary. USEPA Method 1623.1 is a widely used and validated protocol for detecting these parasites in water samples. It consists of a concentration step, followed by parasite purification and visualization by immunofluorescence microscopy. Although efficient, this method has a high cost particularly due to the immunomagnetic separation (IMS) step, which is most needed with complex and highly contaminated samples. Based on this, the present study aimed to determine whether it is possible to maintain the efficiency of Method 1623.1 while reducing the amount of beads per reaction, using as a matrix the challenge water recommended by the World Health Organization. As for Giardia cysts, a satisfactory recovery efficiency (RE) was obtained using 50% less IMS beads. This was evaluated both with a commercial cyst suspension (56.1% recovery) and an analytical quality assessment (47.5% recovery). Although RE rates obtained for Cryptosporidium parvum did not meet Method 1623.1 criteria in any of the experimental conditions tested, results presented in this paper indicated the relevance of the described adaptations, even in challenge water.

Ex vivo fecal fermentation of human ileal fluid collected after raspberry consumption modifies (poly)phenolics and modulates genoprotective effects in colonic epithelial cells.

Diets rich in fruit and vegetables are associated with a decreased incidence of colorectal cancer (CRC) due, in part, to the bioactive (poly)phenolic components and their microbiota-mediated metabolites. This study investigated how such compounds, derived from ingested raspberries in the gastrointestinal tract, may exert protective effects by reducing DNA damage. Ileal fluids collected pre- and post-consumption of 300 g of raspberries by ileostomists (n = 11) were subjected to 24 h ex vivo fermentation with fecal inoculum to simulate interaction with colonic microbiota. The impact of fermentation on (poly)phenolics in ileal fluid was determined and the bioactivity of ileal fluids pre- and post fermentation investigated. (Poly)phenolic compounds including sanguiin H-6, sanguiin H-10 and cyanidin-3-O-sophoroside decreased significantly during fermentation while, in contrast, microbial catabolites, including 3-(3'-hydroxyphenyl)propanoic acid, 3-hydroxybenzoic acid and benzoic acid increased significantly. The post-raspberry ileal fermentate from 9 of the 11 ileostomates significantly decreased DNA damage (~30%) in the CCD 841 CoN normal cell line using an oxidative challenge COMET assay. The raspberry ileal fermentates also modulated gene expression of the nuclear factor 2-antioxidant responsive element (Nrf2-ARE) pathway involved in oxidative stress cytoprotection, namely Nrf2, NAD(P)H dehydrogenase, quinone-1 and heme oxygenase-1. Four of the phenolic catabolites were assessed individually, each significantly reducing DNA damage from an oxidative challenge over a physiologically relevant 10-100 µM range. They also induced a differential pattern of expression of key genes in the Nrf2-ARE pathway in CCD 841 CoN cells. The study indicates that the colon-available raspberry (poly)phenols and their microbial-derived catabolites may play a role in protection against CRC in vivo.

Competent but complex communication: The phenomena of pheromone-responsive plasmids.

Enterococci are robust gram-positive bacteria that are found in a variety of surroundings and that cause a significant number of healthcare-associated infections. The genus possesses a high-efficiency pheromone-responsive plasmid (PRP) transfer system for genetic exchange that allows antimicrobial-resistance determinants to spread within bacterial populations. The pCF10 plasmid system is the best characterised, and although other PRP systems are structurally similar, they lack exact functional homologues of pCF10-encoded genes. In this review, we provide an overview of the enterococcal PRP systems, incorporating functional details for the less-well-defined systems. We catalogue the virulence-associated elements of the PRPs that have been identified to date, and we argue that this reinforces the requirement for elucidation of the less studied systems.

Furanone quorum-sensing inhibitors with potential as novel therapeutics against Pseudomonas aeruginosa.

Micro-organisms use quorum sensing (QS), a cell density-dependent process, to communicate. This QS mode of interchange leads to the production of a variety of virulence factors, co-ordination of complex bacterial behaviours, such as swarming motility, degradation of host tissue and biofilm formation. QS is implicated in numerous human infections and consequently researchers have sought ways of effectively inhibiting the process in pathogenic bacteria. Two decades ago, furanones were the first class of chemical compounds identified as Pseudomonas aeruginosa QS inhibitors (QSIs). P. aeruginosa is a ubiquitous organism, capable of causing a wide range of infections in humans, including eye and ear infections, wound infections and potentially fatal bacteraemia and thus novel treatments against this organism are greatly needed. This review provides a brief background on QS and the use of furanones as QSIs. Based on the effectiveness of action, both in vivo and in vitro, we will explore the use of furanones as potential antimicrobial therapeutics and conclude with open questions.

Modest improvement in CVD risk markers in older adults following quinoa (Chenopodium quinoa Willd.) consumption: a randomized-controlled crossover study with a novel food product.

PURPOSE: To investigate the effect of consuming quinoa biscuits on markers of CVD risk over 4 weeks in free-living older adults. METHODS: A randomized-controlled, double-blind crossover trial was conducted in which consenting healthy adults aged 50-75 years (n = 40) consumed 15 g quinoa biscuits (60 g quinoa flour/100 g) or control iso-energetic biscuits (made using wheat flour) daily for 28 consecutive days (4 weeks), in addition to their normal diet. Following a 6-week washout, participants consumed the alternate biscuit for a final 4 weeks. Anthropometry and fasted blood samples were obtained before and after each intervention period. RESULTS: At the beginning of the trial, mean ± SD total cholesterol concentrations were 6.02 ± 1.22 mmol/L (3.7-9.2 mmol/L); 33 participants (82.5%) had high cholesterol (> 5 mmol/L). No participants were lost to follow-up and there were no changes in habitual dietary intakes or levels of physical activity between each 4-week intervention period. Significantly greater decreases in total and LDL cholesterol concentrations (- 0.30 ± 0.58 and - 0.25 ± 0.38 mmol/L, respectively), TC: HDL ratio (- 0.11 ± 0.30), weight (- 0.61 ± 0.89 kg) and BMI (- 0.22 ± 0.34 kg/m2) were apparent following consumption of the quinoa versus control biscuits (all P < 0.05). Changes in triglycerides, HDL cholesterol, or PUFA or CRP concentrations were not significant between treatment groups. CONCLUSION: Consumption of novel quinoa biscuits produced small, but favorable changes in body weight, BMI, and circulating cholesterol concentrations, all of which may contribute to lowered CVD risk in older adults.

Electroanalytical properties of chlorophenol red at disposable carbon electrodes: Implications for Escherichia coli detection.

The use of coliforms and Escherichia coli as indicator species for assessing the quality of water is well established and a large variety of methods based on ß-galactosidase (B-GAL) activity, inherent to the microbes within this classification, have arisen to enable their detection and enumeration. Chlorophenol red (CPR) is widely used as a chromogenic label, but its capacity for translation to electroanalytical devices has yet to be fully explored. The CPR moiety is capable of undergoing oxidation at carbon substrates (+0.7 V) giving rise to a variety of phenolic intermediates. Electrochemical, XPS and enzymatic techniques were employed to characterise the underpinning chemistry and the intermediate identified as a 1,2-quinone derivative in which the chlorine substituent is retained. The latter was found to accumulate at the electrode and, in contrast to the parent CPR, was found to be detected at a significantly less positive potential (+0.3 V). Bacterial hydrolysis of a CPR labelled substrate was demonstrated with the 1,2-quinone oxidation product found to accumulate at the electrode and detected using square wave voltammetry. Proof of concept for the efficacy of the alternative electrode pathway was established through the detection of E.coli after an incubation time of 2.5 h with no interference from the labelled substrates.

Increased sporulation underpins adaptation of Clostridium difficile strain 630 to a biologically-relevant faecal environment, with implications for pathogenicity.

Clostridium difficile virulence is driven primarily by the processes of toxinogenesis and sporulation, however many in vitro experimental systems for studying C. difficile physiology have arguably limited relevance to the human colonic environment. We therefore created a more physiologically-relevant model of the colonic milieu to study gut pathogen biology, incorporating human faecal water (FW) into growth media and assessing the physiological effects of this on C. difficile strain 630. We identified a novel set of C. difficile-derived metabolites in culture supernatants, including hexanoyl- and pentanoyl-amino acid derivatives by LC-MSn. Growth of C. difficile strain 630 in FW media resulted in increased cell length without altering growth rate and RNA sequencing identified 889 transcripts as differentially expressed (p < 0.001). Significantly, up to 300-fold increases in the expression of sporulation-associated genes were observed in FW media-grown cells, along with reductions in motility and toxin genes' expression. Moreover, the expression of classical stress-response genes did not change, showing that C. difficile is well-adapted to this faecal milieu. Using our novel approach we have shown that interaction with FW causes fundamental changes in C. difficile biology that will lead to increased disease transmissibility.

Impact of a (poly)phenol-rich extract from the brown algae Ascophyllum nodosum on DNA damage and antioxidant activity in an overweight or obese population: a randomized controlled trial.

Background: Epidemiologic evidence suggests that a diet rich in (poly)phenols has beneficial effects on many chronic diseases. Brown seaweed is a rich source of (poly)phenols. Objective: The aim of this study was to investigate the bioavailability and effect of a brown seaweed (Ascophyllum nodosum) (poly)phenol extract on DNA damage, oxidative stress, and inflammation in vivo. Design: A randomized, double-blind, placebo-controlled crossover trial was conducted in 80 participants aged 30-65 y with a body mass index (in kg/m2) ≥25. The participants consumed either a 400-mg capsule containing 100 mg seaweed (poly)phenol and 300 mg maltodextrin or a 400-mg maltodextrin placebo control capsule daily for an 8-wk period. Bioactivity was assessed with a panel of blood-based markers including lymphocyte DNA damage, plasma oxidant capacity, C-reactive protein (CRP), and inflammatory cytokines. To explore the bioavailability of seaweed phenolics, an untargeted metabolomics analysis of urine and plasma samples after seaweed consumption was determined by ultra-high-performance liquid chromatography-high-resolution mass spectrometry. Results: Consumption of the seaweed (poly)phenols resulted in a modest decrease in DNA damage but only in a subset of the total population who were obese. There were no significant changes in CRP, antioxidant status, or inflammatory cytokines. We identified phlorotannin metabolites that are considered potential biomarkers of seaweed consumption including pyrogallol/phloroglucinol-sulfate, hydroxytrifurahol A-glucuronide, dioxinodehydroeckol-glucuronide, diphlorethol sulfates, C-O-C dimer of phloroglucinol sulfate, and C-O-C dimer of phloroglucinol. Conclusions: To the best of our knowledge, this work represents the first comprehensive study investigating the bioactivity and bioavailability of seaweed (poly)phenolics in human participants. We identified several potential biomarkers of seaweed consumption. Intriguingly, the modest improvements in DNA damage were observed only in the obese subset of the total population. The subgroup analysis should be considered exploratory because it was not preplanned; therefore, it was not powered adequately. Elucidation of the biology underpinning this observation will require participant stratification according to weight in future studies. This trial was registered at clinicaltrials.gov as NCT02295878.

Inactivation of the dnaK gene in Clostridium difficile 630 Δerm yields a temperature-sensitive phenotype and increases biofilm-forming ability.

Clostridium difficile infection is a growing problem in healthcare settings worldwide and results in a considerable socioeconomic impact. New hypervirulent strains and acquisition of antibiotic resistance exacerbates pathogenesis; however, the survival strategy of C. difficile in the challenging gut environment still remains incompletely understood. We previously reported that clinically relevant heat-stress (37-41 °C) resulted in a classical heat-stress response with up-regulation of cellular chaperones. We used ClosTron to construct an insertional mutation in the dnaK gene of C. difficile 630 Δerm. The dnaK mutant exhibited temperature sensitivity, grew more slowly than C. difficile 630 Δerm and was less thermotolerant. Furthermore, the mutant was non-motile, had 4-fold lower expression of the fliC gene and lacked flagella on the cell surface. Mutant cells were some 50% longer than parental strain cells, and at optimal growth temperatures, they exhibited a 4-fold increase in the expression of class I chaperone genes including GroEL and GroES. Increased chaperone expression, in addition to the non-flagellated phenotype of the mutant, may account for the increased biofilm formation observed. Overall, the phenotype resulting from dnaK disruption is more akin to that observed in Escherichia coli dnaK mutants, rather than those in the Gram-positive model organism Bacillus subtilis.

The composition of potentially bioactive triterpenoid glycosides in red raspberry is influenced by tissue, extraction procedure and genotype.

The beneficial effects of consumption of berry fruits on a range of chronic diseases has been attributed (at least in part) to the presence of unique phytochemicals. Recently, we identified novel ursolic acid-based triterpenoid glycosides (TTPNs) in raspberry fruit and demonstrated their survival in human ileal fluids after feeding which confirmed their colon-availability in vivo. In this paper, in vitro digestion studies demonstrated that certain TTPNs were stable under gastrointestinal conditions and confirmed that these components may have been responsible for bioactivity noted in previous studies. Sequential extractions of raspberry puree, isolated seeds and unseeded puree showed that certain TTPN components (e.g. peak T1 m/z 679, and T2 m/z 1358) had different extractabilities in water/solvent mixes and were differentially associated with the seeds. Purified seed TTPNs (mainly T1 and T2) were shown to be anti-genotoxic in HT29 and CCD841 cell based in vitro colonocyte models. Further work confirmed that the seeds contained a wider range of TTPN-like components which were also differentially extractable in water/solvent mixes. This differential extractability could influence the TTPN composition and potential bioactivity of the extracts. There was considerable variation in total content of TTPNs (∼3-fold) and TTPN composition across 13 Rubus genotypes. Thus, TTPNs are likely to be present in raspberry juices and common extracts used for bioactivity studies and substantial variation exists in both content and composition due to genetics, tissue source or extraction conditions, which may all affect observed bioactivity.