Search

Pilling is a form of textile mechanical damage, forming fibrous bobbles on the surface of garments, resulting in premature disposal of clothing by consumers. However, our understanding on how the structural properties of the cellulosic matrix compliment the three-dimensional shape of cotton pills remains limited. This knowledge gap has hindered the development of effective 'pillase' technologies over the past 20 years due to challenges in balancing depilling efficacy with fabric integrity preservation. Therefore, the main focus here was characterising the role of cellulose and the hemicellulose components in cotton textiles to elucidate subtle differences between the chemistry of pills and fibre regions involved in structural integrity. State-of-the-art bioimaging using carbohydrate binding modules, monoclonal antibodies, and Leica SP8 and a Nikon A1R confocal microscopes, revealed the biophysical structure of cotton pills for the first time. Identifying regions of increased crystalline cellulose in the base of anchor fibres and weaker amorphous cellulose at dislocations in their centres, enhancing our understanding of current enzyme specificity. Surprisingly, pills contained a 7-fold increase in the concentration of xyloglucan compared to the main textile. Therefore, xyloglucan offers a previously undescribed target for overcoming this benefit-to-risk paradigm, suggesting a role for xyloglucanase enzymes in future pillase systems., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Neil Lant reports financial support was provided by Procter and Gamble. Neil Lant reports a relationship with Procter and Gamble that includes: employment. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Marine gel particles (MGP) are amorphous hydrogel exudates from bacteria and microalgae that are ubiquitous in the oceans, but their biochemical composition and function are poorly understood. While dynamic ecological interactions between marine microorganisms and MGPs may result in the secretion and mixing of bacterial extracellular polymeric substances (EPS) such as nucleic acids, compositional studies currently are limited to the identification of acidic polysaccharides and proteins in transparent exopolymer particles (TEP) and Coomassie stainable particles (CSP). Previous studies targeted MGPs isolated by filtration. We developed a new way of isolating MGPs from seawater in liquid suspension and applied it to identify extracellular DNA (eDNA) in North Sea surface seawater. Seawater was filtered onto polycarbonate (PC) filters with gentle vacuum filtration, and then the filtered particles were gently resuspended in a smaller volume of sterile seawater. The resulting MGPs ranged in size from 0.4 to 100 µm in diameter. eDNA was detected by fluorescent microscopy using YOYO-1 (for eDNA), with Nile red (targeting cell membranes) as a counterstain. TOTO-3 was also used to stain eDNA, with ConA to localise glycoproteins and SYTO-9 for the live/dead staining of cells. Confocal laser scanning microscopy (CLSM) revealed the presence of proteins and polysaccharides. We found eDNA to be universally associated with MGPs. To further elucidate the role of eDNA, we established a model experimental MGP system using bacterial EPS from Pseudoalteromonas atlantica that also contained eDNA. Our results clearly demonstrate the occurrence of eDNA in MGPs, and should aid furthering our understanding of the micro-scale dynamics and fate of MGPs that underly the large-scale processes of carbon cycling and sedimentation in the ocean.

Background: We convened a two-round, modified Delphi panel to identify and reach consensus on additional potential quality indicators (QIs) for nursing home residents with dementia., Methods: The study team identified 12 potential QIs for nursing home dementia care and treatment of behavioral disturbances based on review of the literature. All proposed QIs were readily available in administrative claims data. Panelists rated each QI on importance, usefulness, and feasibility (a total of 36 items) using a 9-point Likert scale. Data were collected using an online survey platform and virtual group discussion. We defined consensus as ≥70% of the panelists responding within a three-point range surrounding the median. A QI achieved relevance on a domain (importance, usefulness, feasibility) when the panel reached consensus and a median rating of 7-9., Results: The study had a 100% response rate for both survey rounds. Twenty-four items achieved consensus, with 15 reaching relevance with a median >7. Three QIs (percent of long-stay residents with dementia prescribed APs, percent with physical restraint use, and percent with a positive behavioral symptom score) reached consensus at the highest median score (9) for importance. Only 2 of the 12 proposed QIs reached relevance on all three domains: percent of long-stay residents with dementia prescribed antipsychotics (APs) and percent prescribed benzodiazepines., Conclusions: Of the proposed QIs, our panel of dementia care experts only reached consensus on two QIs: measuring long-stay resident prescriptions of APs and benzodiazepines. Challenges remain in identifying QIs that meet threshold of all three areas and accurately reflect quality nursing home dementia care.

Plastic pollution has now been found within multiple ecosystems across the globe. Characterisation of microbial assemblages associated with marine plastic, or the so-called 'plastisphere', has focused predominantly on plastic in the epipelagic zone. Whether this community includes taxa that are consistently enriched on plastic compared to surrounding non plastic surfaces is unresolved, as are the ecological implications. The deep sea is likely a final sink for most of the plastic entering the ocean, yet there is limited information on microbial colonisation of plastic at depth. The aim of this study was to investigate deep-sea microbial communities associated with polystyrene (PS) and polyurethane (PU) with Bath stone used as a control. The substrates (n = 15) were deployed in the Rockall Trough (Atlantic), and recovered 420 days later from a depth of 1796 m. To characterise the bacterial communities, 16S rRNA genes were sequenced using the Illumina MiSeq platform. A dominant core microbiome (taxa shared across all substrates) comprised 8% of total ASVs (amplicon sequence variant) and accounted for 92% of the total community reads. This suggests that many commonly reported members of the plastisphere are simply opportunistic which freely colonise any hard surface. Transiently associated species consisted of approximately 7% of the total community. Thirty genera were enriched on plastic (P < 0.05), representing 1% of the total community. The discovery of novel deep-sea enriched taxa included Aurantivirga, Algivirga, IheB3-7, Spirosoma, HTCC5015, Ekhidna and Calorithrix on PS and Candidatus Obscuribacter, Haloferula, Marine Methylotrophic Group 3, Aliivibrio, Tibeticola and Dethiosulfatarculus on PU. This small fraction of the microbiome include taxa with unique metabolic abilities and show how bacterial communities can be shaped by plastic pollution at depth. This study outlines a novel approach in categorising the plastisphere to elucidate the ecological implications of enriched taxa that show an affinity for colonising plastic., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Epibiotic bacteria associated with the filamentous marine cyanobacterium Moorea producens were explored as a novel source of antibiotics and to establish whether they can produce cyclodepsipeptides on their own. Here, we report the isolation of micrococcin P1 ( 1 ) (C 48 H 49 N 13 O 9 S 6 ; obs. m / z 1144.21930/572.60381) and micrococcin P2 ( 2 ) (C 48 H 47 N 13 O 9 S 6 ; obs. m / z 1142.20446/571.60370) from a strain of Bacillus marisflavi isolated from M. producens ' filaments. Interestingly, most bacteria isolated from M. producens ' filaments were found to be human pathogens. Stalked diatoms on the filaments suggested a possible terrestrial origin of some epibionts. CuSO 4 ·5H 2 O assisted differential genomic DNA isolation and phylogenetic analysis showed that a Kenyan strain of M. producens differed from L. majuscula strain CCAP 1446/4 and L. majuscula clones. Organic extracts of the epibiotic bacteria Pseudoalteromonas carrageenovora and Ochrobactrum anthropi did not produce cyclodepsipeptides. Further characterization of 24 Firmicutes strains from M. producens identified extracts of B. marisflavi as most active. Our results showed that the genetic basis for synthesizing micrococcin P1 ( 1 ), discovered in Bacillus cereus ATCC 14579, is species/strain-dependent and this reinforces the need for molecular identification of M. producens species worldwide and their epibionts. These findings indicate that M. producens -associated bacteria are an overlooked source of antimicrobial compounds.

Bacterial biofilms in natural and artificial environments perform a wide array of beneficial or detrimental functions and exhibit resistance to physical as well as chemical perturbations. In dynamic environments, where periodic or aperiodic flows over surfaces are involved, biofilms can be subjected to large shear forces. The ability to withstand these forces, which is often attributed to the resilience of the extracellular matrix. This attribute of the extracellular matrix is referred to as viscoelasticity and is a result of self-assembly and cross-linking of multiple polymeric components that are secreted by the microbes. We aim to understand the viscoelastic characteristic of biofilms subjected to large shear forces by performing Large Amplitude Oscillatory Shear (LAOS) experiments on four species of bacterial biofilms: Bacillus subtilis, Comamonas denitrificans, Pseudomonas fluorescens and Pseudomonas aeruginosa. We find that nonlinear viscoelastic measures such as intracycle strain stiffening and intracycle shear thickening for each of the tested species, exhibit subtle or distinct differences in the plot of strain amplitude versus frequency (Pipkin diagram). The biofilms also exhibit variability in the onset of nonlinear behaviour and energy dissipation characteristics, which could be a result of heterogeneity of the extracellular matrix constituents of the different biofilms. The results provide insight into the nonlinear rheological behaviour of biofilms as they are subjected to large strains or strain rates; a situation that is commonly encountered in nature, but rarely investigated.

The secretion of extracellular polymeric substances provides an evolutionary advantage found in many organisms that can adhere to surfaces and cover themselves in a protective matrix. This ability is found in prokaryotes, archaea and eukaryotes, all of which use functionally similar polysaccharides, proteins and nucleic acids to form extracellular matrices, mucus and bioadhesive substances. These macromolecules have been investigated from the perspective of polymer biophysics, and theories to help understand adhesion, viscosity and gelling have been developed. These properties can be measured experimentally using straightforward methods such as cell counting as well as more advanced techniques such as atomic force microscopy and rheometry. An integrated understanding of the properties and uses of adhesive macromolecules across kingdoms is also important and can provide the basis for a range of biotechnological and medical applications. This article is part of the theme issue 'Transdisciplinary approaches to the study of adhesion and adhesives in biological systems'.

The influence of laundry washing parameters on the release of microfibers (MF) from polyester textiles was studied. These fibers are an important type of microplastic pollution. However, the factors which affect MF release during laundry are poorly understood and more rigorous methods for quantifying this release are needed. A novel method was therefore developed using a tergotometer with eight 1000 mL washing vessels and the CIELab color space measure of lightness ( L *). L * was related to the mass of released MFs by creating a calibration curve to quantify the amounts of MFs released from textiles during washing. This method was used to investigate the effect of water-volume, agitation, temperature, and duration of the wash on MF release. Counterintuitively, increased water-volume, characteristic of European "delicate" cycles, resulted in the greatest release of MFs. Full-scale testing was then carried out using domestic washing machines with real consumer cycles to determine the effect of cycle type on MF release. In the first wash, delicate wash cycles released 800 000 more MFs (94 mg/kg) per wash than a lower water-volume standard wash and also increased MF release in subsequent washing cycles ( P < 0.05). These results indicate that a high water-volume-to-fabric ratio is the most influential factor for MF release, rather than agitation as previously thought. Therefore, consumers can reduce MF release by avoiding high water-volume washes (delicate cycles), transitioning to appliances that use a lower water-volume (North American high-efficiency washing machines), and ensuring that full wash loads are used.

Most marine benthic invertebrates have a pelagic larval phase, after which they settle preferentially on or near conspecific adults, forming aggregations. Although settlement pheromones from conspecific adults have been implicated as critical drivers of aggregation for more than 30 years, surprisingly few have been unambiguously identified. Here we show that in the invasive dreissenid mussel Mytilopsis sallei (an ecological and economic pest), three common purines (adenosine, inosine, and hypoxanthine) released from adults in a synergistic and precise ratio (1:1.125:3.25) serve as an aggregation pheromone by inducing conspecific larval settlement and metamorphosis. Our results demonstrate that simple common metabolites can function as species-specific pheromones when present in precise combinations. This study provides important insights into our understanding of the ecology and communication processes of invasive organisms and indicates that the combination and ratio of purines might be critical for purine-based signaling systems that are fundamental and widespread in nature., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

Biofilms occur in a broad range of environments under heterogeneous physicochemical conditions, such as in bioremediation plants, on surfaces of biomedical implants, and in the lungs of cystic fibrosis patients. In these scenarios, biofilms are subjected to shear forces, but the mechanical integrity of these aggregates often prevents their disruption or dispersal. Biofilms' physical robustness is the result of the multiple biopolymers secreted by constituent microbial cells which are also responsible for numerous biological functions. A better understanding of the role of these biopolymers and their response to dynamic forces is therefore crucial for understanding the interplay between biofilm structure and function. In this paper, we review experimental techniques in rheology, which help quantify the viscoelasticity of biofilms, and modeling approaches from soft matter physics that can assist our understanding of the rheological properties. We describe how these methods could be combined with synthetic biology approaches to control and investigate the effects of secreted polymers on the physical properties of biofilms. We argue that without an integrated approach of the three disciplines, the links between genetics, composition, and interaction of matrix biopolymers and the viscoelastic properties of biofilms will be much harder to uncover., (Copyright © 2019 Charlton et al.)

The vast majority of bacteria present in the natural environment are present in the form of aggregates and/or biofilms. Microbial aggregates are ubiquitous in the marine environment and are inhabited by diverse microbial communities which often express intense extracellular enzymatic activities. However, the secretion of an important group of enzymes, DNases, by bacteria from marine aggregates has not been studied, despite the importance of these aggregates in biogeochemical cycling of nutrients in the oceans. In this work, we therefore, employed both culture-based and bioinformatics approaches to understand the diversity of bacterial DNases in marine bacterioplankton. We found that 34% of 345 strains of attached and non-attached marine bacteria showed extracellular DNase activity. Most of these isolates belong to Proteobacteria (53%) and Firmicutes (34%). Secretion of DNases by bacteria isolated from marine gel particles (MGP) is reported here for the first time. Then, to further understand the wider diversity of the potential to produce DNases, sequences were compared using 2316 whole genome and 42 metagenome datasets. Thirty-nine different taxonomic groups corresponding to 10 bacterial phyla were found to encode genes responsible for DNase secretion. This study highlights the unexpected and widespread presence of DNase secretion in bacteria in general and in MGP more specifically. This has important implications for understanding the dynamics and fate of marine microbial aggregates in the oceans.

UV resistance of bacteria isolated from the marine cyanobacterium Moorea producens has not been observed previously, findings which highlight how unsafe germicidal UV irradiation for sterilization of air, food, and water could be. Further, UV resistance of Bacillus licheniformis is being observed for the first time. This study focused on bacteria isolated from the marine cyanobacterium M. producens collected off the Kenyan coast at Shimoni, Wasini, Kilifi, and Mida. UV irradiance of isolates (302 nm, 70 W/m 2 , 0-1 hr) established B. licheniformis as the most UV resistant strain, with the following order of taxon resistance: Bacilli> γ proteobacteria > Actinobacteria. UV resistance was independent of pigmentation. The maximum likelihood phylogenetic distance determined for both B. licheniformis and Bacillus aerius relative to M. producens CCAP 1446/4 was 2.0. Survival of B. licheniformis upon UV irradiance followed first-order kinetics (k = 0.035/min, R 2  = 0.88). Addition of aqueous extracts (2, 10, 20 and 40 mg/ml) of this B. licheniformis strain on the less resistant Marinobacterium stanieri was not significant, however, the commercial sunscreen benzophenone-3 (BP-3) positive control and the time of irradiance were significant. Detection of bacteria on M. producens filaments stained with acridine orange confirmed its nonaxenic nature. Although the chemistry of UV resistance in cyanobacteria has been studied in depth revealing for example the role of mycosporine like amino acids (MAAs) in UV resistance less is known about how bacteria resist UV irradiation. This is of interest since cyanobacteria live in association with bacteria., (© 2018 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Mucus layers often provide a unique and multi-functional hydrogel interface between the epithelial cells of organisms and their external environment. Mucus has exceptional properties including elasticity, changeable rheology and an ability to self-repair by re-annealing, and is therefore an ideal medium for trapping and immobilising pathogens and serving as a barrier to microbial infection. The ability to produce a functional surface mucosa was an important evolutionary step, which evolved first in the Cnidaria, which includes corals, and the Ctenophora. This allowed the exclusion of non-commensal microbes and the subsequent development of the mucus-lined digestive cavity seen in higher metazoans. The fundamental architecture of the constituent glycoprotein mucins is also evolutionarily conserved. Although an understanding of the biochemical interactions between bacteria and the mucus layer are important to the goal of developing new antimicrobial strategies, they remain relatively poorly understood. This review summarises the physicochemical properties and evolutionary importance of mucus, which make it so successful in the prevention of bacterial infection. In addition, the strategies developed by bacteria to counteract the mucus layer are also explored., Competing Interests: The authors declare no competing interests.

The marine environment represents an underexploited resource for the discovery of novel products, despite its high level of biological and chemical diversity. With increasing awareness of the harmful effects of chronic ultraviolet exposure, and a universal desire to improve cosmetic appearance, the market for new cosmetic ingredients is growing, and current trends have generated a greater demand for products sourced from the environment. A growing number of novel molecules from marine flora and fauna exhibit potent and effective dermatological activities. Secondary metabolites isolated from macroalgae, including carotenoids and polyphenols, have demonstrated antioxidant, anti-ageing and anti-inflammatory activities. In addition, marine extremophilic bacteria have recently been shown to produce bioactive exopolymeric molecules, some of which have been commercialized. Available data on their activities show significant antioxidant, moisturizing and anti-ageing activities, but a more focussed investigation into their mechanisms and applications is required. This review surveys the reported biological activities of an emerging and growing portfolio of marine molecules that show promise in the treatment of cosmetic skin problems including ultraviolet damage, ageing and cutaneous dryness., (© 2017 Society of Cosmetic Scientists and the Société Française de Cosmétologie.)

Bacterial biofilms are a complex architecture of cells that grow on moist interfaces, and are held together by a molecular glue of extracellular proteins, sugars and nucleic acids. Biofilms are particularly problematic in human healthcare as they can coat medical implants and are thus a potential source of disease. The enzymatic dispersal of biofilms is increasingly being developed as a new strategy to treat this problem. Here, we have characterized NucB, a biofilm-dispersing nuclease from a marine strain of Bacillus licheniformis, and present its crystal structure together with the biochemistry and a mutational analysis required to confirm its active site. Taken together, these data support the categorization of NucB into a unique subfamily of the ββα metal-dependent non-specific endonucleases. Understanding the structure and function of NucB will facilitate its future development into an anti-biofilm therapeutic agent., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Omega-3 fatty acids are products of secondary metabolism, essential for growth and important for human health. Although there are numerous reports of bacterial production of omega-3 fatty acids, less information is available on the biotechnological production of these compounds from bacteria. The production of eicosapentaenoic acid (EPA, 20:5ω3) by a new species of marine bacteria Shewanella electrodiphila MAR441T was investigated under different fermentation conditions. This strain produced a high percentage (up to 26%) of total fatty acids and high yields (mg / g of biomass) of EPA at or below the optimal growth temperature. At higher growth temperatures these values decreased greatly. The amount of EPA produced was affected by the carbon source, which also influenced fatty acid composition. This strain required Na+ for growth and EPA synthesis and cells harvested at late exponential or early stationary phase had a higher EPA content. Both the highest amounts (20 mg g-1) and highest percent EPA content (18%) occurred with growth on L-proline and (NH4)2SO4. The addition of cerulenin further enhanced EPA production to 30 mg g-1. Chemical mutagenesis using NTG allowed the isolation of mutants with improved levels of EPA content (from 9.7 to 15.8 mg g-1) when grown at 15°C. Thus, the yields of EPA could be substantially enhanced without the need for recombinant DNA technology, often a commercial requirement for food supplement manufacture.

Marine biofouling is a worldwide problem affecting maritime industries. Global concerns about the high toxicity of antifouling paints have highlighted the need to develop less toxic antifouling coatings. Chitosan is a natural polymer with antimicrobial, antifungal and antialgal properties that is obtained from partial deacetylation of crustacean waste. In the present study, nanocomposite chitosan-zinc oxide (chitosan-ZnO) nanoparticle hybrid coatings were developed and their antifouling activity was tested. Chitosan-ZnO nanoparticle coatings showed anti-diatom activity against Navicula sp. and antibacterial activity against the marine bacterium Pseudoalteromonas nigrifaciens. Additional antifouling properties of the coatings were investigated in a mesocosm study using tanks containing natural sea water under controlled laboratory conditions. Each week for four weeks, biofilm was removed and analysed by flow cytometry to estimate total bacterial densities on the coated substrates. Chitosan-ZnO hybrid coatings led to better inhibition of bacterial growth in comparison to chitosan coatings alone, as determined by flow cytometry. This study demonstrates the antifouling potential of chitosan-ZnO nanocomposite hybrid coatings, which can be used for the prevention of biofouling., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Strains MAR441(T) and MAR445 were isolated from Mid-Atlantic Ridge sediments from a depth of 2734 m, and were found to belong to the genus Shewanella. The strains were rod-shaped, pigmented, non-motile and capable of anaerobic growth either by fermentation of carbohydrates or by anaerobic respiration. The strains utilized a variety of electron acceptors, including nitrate and ferric compounds, and could utilize peptone when grown anaerobically in a two-chambered microbial fuel cell, which used carbon cloth electrodes and delivered a stable power output of ,150-200 mW m(-2). The major fatty acids were typical of the genus Shewanella, with major components C13 : 0, iso-C13 : 0, iso-C15 : 0, C16 : 0, C16 : 1ω7c, C18 : 1ω7c and C20 : 5ω3 fatty acids. The DNA G+C content of strains MAR441(T) and MAR445 was 42.4 mol%. 16S rRNA gene sequence analysis indicated that strains MAR441(T) and MAR445 were most closely related to Shewanella olleyana (sequence similarities 97.9% to the type strain). DNA-DNA hybridization demonstrated only 15.6-37.2% relatedness between strain MAR441(T) and the type strains of related species of the genus Shewanella. Phenotypic characteristics confirmed that these isolates constituted a novel species of the genus Shewanella, for which the name Shewanella electrodiphila sp. nov. is proposed; the type strain is MAR441(T) (5ATCC BAA-2408(T) = DSM 24955(T)).

The extracellular matrix of microbial biofilms is critical for surface adhesion and nutrient homeostasis. Evidence is accumulating that extracellular DNA plays a number of important roles in biofilm integrity and formation on hard and soft tissues in the oral cavity. Here, we summarise recent developments in the field and consider the potential of targeting DNA for oral biofilm control., (Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Strains MAR441T and MAR445 were isolated from Mid-Atlantic Ridge (MAR) sediments with a depth of 2,734 m, and found to be member of the genus Shewanella. The strains were rod shaped, pigmented, non-motile, and capable of anaerobic growth either by fermentation of carbohydrates or by anaerobic respiration. The strains utilized a variety of electron acceptors, including nitrate and ferric compounds and could utilize peptone when grown anaerobically in a two-chambered microbial fuel cell (MFC), which use carbon cloth electrodes and deliver a stable power output of ~150-200 mW/m2. The major fatty acids were typical of the genus Shewanella, with major components of C13:0, iso-C13:0, iso-C15:0, C16:0, C16:1ω7c, C18:1ω7c and C20:5ω3 fatty acids. The DNA G+C content of strains MAR441T and MAR445 were 42.4 mol %. 16S rRNA gene sequence analysis indicated that strains MAR441T and MAR445 were most closely related to Shewanella olleyana (sequence similarities 97.9 %). DNA-DNA hybridization demonstrated only 15.6-37.2 % relatedness between strain MAR441T and the type strains of related Shewanella species. Phenotypic characteristics confirmed that these isolates constituted a novel species of the genus Shewanella. The type strain of Shewanella dovemarinensis is MAR441T (=ATCC BAA-2408 T =DSM24955 T)., (Copyright © 2014, the Society for General Microbiology.)

The death and lysis of microbial cells leads to the release of cytoplasmic contents, many of which are rapidly degraded by enzymes. However, some macromolecules survive intact and find new functions in the extracellular environment. There is now strong evidence that DNA released from cells during lysis, or sometimes by active secretion, becomes a key component of the macromolecular scaffold in many different biofilms. Enzymatic degradation of extracellular DNA can weaken the biofilm structure and release microbial cells from the surface. Many bacteria produce extracellular deoxyribonuclease (DNase) enzymes that are apparently tightly regulated to avoid excessive degradation of the biofilm matrix. Interfering with these control mechanisms, or adding exogenous DNases, could prove a potent strategy for controlling biofilm growth., (© 2013 The Society for Applied Microbiology.)

The extracellular nuclease, NucB, from Bacillus licheniformis, can digest extracellular DNA in biofilms, causing biofilm dispersal, and may therefore be used commercially to remove biofilms. However, producing quantities of this secreted peptide is difficult and our aim was therefore to improve its laboratory scale production. This study builds on our understanding of B. licheniformis physiology to enhance NucB production. The addition of manganese, which triggers sporulation and enhances NucB expression, lead to a 5-fold increase in NucB production. Optimisation via Placket-Burman design of experiments identified 3 significant medium components and a subsequent Central Composite Design, to determine the optimum levels of these components, resulted in a 10-fold increase to 471U/ml. The optimal phosphate concentration was less than 0.3mM as this is known to inhibit nuclease production. The use of physiologically relevant information combined with optimisation represents a promising approach to increased enzyme production, which may also be widely applicable., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background: The persistent colonization of paranasal sinus mucosa by microbial biofilms is a major factor in the pathogenesis of chronic rhinosinusitis (CRS). Control of microorganisms within biofilms is hampered by the presence of viscous extracellular polymers of host or microbial origin, including nucleic acids. The aim of this study was to investigate the role of extracellular DNA in biofilm formation by bacteria associated with CRS., Methods/principal Findings: Obstructive mucin was collected from patients during functional endoscopic sinus surgery. Examination of the mucous by transmission electron microscopy revealed an acellular matrix punctuated occasionally with host cells in varying states of degradation. Bacteria were observed in biofilms on mucosal biopsies, and between two and six different species were isolated from each of 20 different patient samples. In total, 16 different bacterial genera were isolated, of which the most commonly identified organisms were coagulase-negative staphylococci, Staphylococcus aureus and α-haemolytic streptococci. Twenty-four fresh clinical isolates were selected for investigation of biofilm formation in vitro using a microplate model system. Biofilms formed by 14 strains, including all 9 extracellular nuclease-producing bacteria, were significantly disrupted by treatment with a novel bacterial deoxyribonuclease, NucB, isolated from a marine strain of Bacillus licheniformis. Extracellular biofilm matrix was observed in untreated samples but not in those treated with NucB and extracellular DNA was purified from in vitro biofilms., Conclusion/significance: Our data demonstrate that bacteria associated with CRS form robust biofilms which can be reduced by treatment with matrix-degrading enzymes such as NucB. The dispersal of bacterial biofilms with NucB may offer an additional therapeutic target for CRS sufferers.

Objective: The growth of biofilms on tracheoesophageal speech valves shortens their life span and produces a reservoir of pathogens that may infect the respiratory tract. The authors have discovered a novel nontoxic deoxyribonuclease, NucB, from a marine isolate of Bacillus licheniformis that is effective at dispersing a variety of mono and mixed-species bacterial biofilms. The aim of this preliminary study was to determine whether NucB could also disrupt and remove mixed-species biofilms from tracheoesophageal speech valves., Study Design: Laboratory-based treatment and analysis of discarded tracheoesophageal speech valves., Setting: University human biology laboratory and the Department of Speech and Language Therapy at a tertiary referral hospital., Subjects and Methods: Seventeen ex vivo tracheoesophageal speech valves fouled with natural human biofilms were collected and divided into 2 equal parts. One half was treated with NucB and the other half with a control buffer solution. Biofilm removal was measured by microscopy and by culture of dispersed biofilm organisms on agar plates., Results: Significantly more organisms were released from biofilms using NucB than with buffer solution alone. On nonselective medium, more organisms were cultured in 11 samples (65%, n = 17, P < .05). Using growth media favoring fungi, more organisms were cultured in 14 samples (82%, n = 17, P < .05)., Conclusion: The nontoxic deoxyribonuclease NucB was effective in releasing more microorganisms from biofilms on tracheoesophageal speech valves. This reflects its potential ability to break up and disperse these biofilms. Future studies will aim to develop NucB as a novel agent to prolong the life span of tracheoesophageal speech valves, thus reducing health care costs.

Photosynthetic microorganisms play crucial roles in aquatic ecosystems and are the major primary producers in global marine ecosystems. The discovery of new bacteria and microalgae that play key roles in CO(2) fixation is hampered by the lack of methods to identify hitherto-unculturable microorganisms. To overcome this problem we studied single microbial cells using stable-isotope probing (SIP) together with resonance Raman (RR) microspectroscopy of carotenoids, the light-absorbing pigments present in most photosynthetic microorganisms. We show that fixation of (13)CO(2) into carotenoids produces a red shift in single-cell RR (SCRR) spectra and that this SCRR-SIP technique is sufficiently sensitive to detect as little as 10% of (13)C incorporation. Mass spectrometry (MS) analysis of labelled cellular proteins verifies that the red shift in carotenoid SCRR spectra acts as a reporter of the (13)C content of single cells. Millisecond Raman imaging of cells in mixed cultures and natural seawater samples was used to identify cells actively fixing CO(2), demonstrating that the SCRR-SIP is a noninvasive method for the rapid and quantitative detection of CO(2) fixation at the single cell level in a microbial community. The SCRR-SIP technique may provide a direct method for screening environmental samples, and could help to reveal the ecophysiology of hitherto-unculturable microorganisms, linking microbial species to their ecological function in the natural environment.

Microbial fuel cells (MFCs) can convert organic compounds directly into electricity by catalytic oxidation, and although MFCs have attracted considerable interest, there is little information on the electricity-generating potential of artificial bacterial biofilms. We have used acetate-fed MFCs inoculated with sediment, with two-chamber bottles and carbon cloth electrodes to deliver a maximum power output of ~175 mW · m(-2) and a stable power output of ~105 mW · m(-2). Power production was by direct transfer of electrons to the anode from bacterial consortia growing on the anode, as confirmed by cyclic voltammetry (CV) and scanning electron microscopy (SEM). Twenty different species (74 strains) of bacteria were isolated from the consortium under anaerobic conditions and cultured in the laboratory, of which 34% were found to be exoelectrogens in single-species studies. Exoelectrogenesis by members of the genera Vibrio , Enterobacter , and Citrobacter and by Bacillus stratosphericus was confirmed, by use of culture-based methods, for the first time. An MFC with a natural bacterial consortium showed higher power densities than those obtained with single strains. In addition, the maximum power output could be further increased to ~200 mW · m(-2) when an artificial consortium consisting of the best 25 exoelectrogenic isolates was used, demonstrating the potential for increased performance and underlying the importance of artificial biofilms for increasing power output.

Marine biotechnology is the industrial, medical or environmental application of biological resources from the sea. Since the marine environment is the most biologically and chemically diverse habitat on the planet, marine biotechnology has, in recent years delivered a growing number of major therapeutic products, industrial and environmental applications and analytical tools. These range from the use of a snail toxin to develop a pain control drug, metabolites from a sea squirt to develop an anti-cancer therapeutic, and marine enzymes to remove bacterial biofilms. In addition, well known and broadly used analytical techniques are derived from marine molecules or enzymes, including green fluorescence protein gene tagging methods and heat resistant polymerases used in the polymerase chain reaction. Advances in bacterial identification, metabolic profiling and physical handling of cells are being revolutionised by techniques such as mass spectrometric analysis of bacterial proteins. Advances in instrumentation and a combination of these physical advances with progress in proteomics and bioinformatics are accelerating our ability to harness biology for commercial gain. Single cell Raman spectroscopy and microfluidics are two emerging techniques which are also discussed elsewhere in this issue. In this review, we provide a brief survey and update of the most powerful and rapidly growing analytical techniques as used in marine biotechnology, together with some promising examples of less well known earlier stage methods which may make a bigger impact in the future., (Copyright © 2012. Published by Elsevier Ltd.)

To evaluate a rapid and cost-effective method for monitoring bacteria in ballast water, several marine bacterial isolates were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Since International Maritime Organization (IMO) regulations are concerned with the unintended transportation of pathogenic bacteria through ballast water, emphasis was placed on detecting species of Vibrio, enterococci and coliforms. Seawater samples collected from the North Sea were incubated in steel ballast tanks and the presence of potentially harmful species of Pseudomonas was also investigated. At the genus-level, the identification of thirty six isolates using MALDI-TOF MS produced similar results to those obtained by 16S rRNA gene sequencing. No pathogenic species were detected either by 16S rRNA gene analysis or by MALDI-TOF MS except for the opportunistically pathogenic bacterium Pseudomonas aeruginosa. In addition, in house software that calculated the correlation coefficient values (CCV) of the mass spectral raw data and their variation was developed and used to allow the rapid and efficient identification of marine bacteria in ballast water for the first time.

Mapping the distribution of phylogenetically distinct bacteria in natural environments is of primary importance to an understanding of ecological dynamics. Here we present a quantitative PCR (qPCR) assay for the analysis of higher taxa composition in natural communities that advances previously available methods by allowing quantification of several taxa during the same qPCR run. Existing primers targeting the 16S rRNA gene specific for Firmicutes, Actinobacteria, Bacteroidetes and for the α and γ subdivisions of the Proteobacteria were improved by largely increasing the coverage of the taxon they target without diminishing their specificity. The qPCR assay was validated in vitro testing artificial mixtures of 16S rRNA sequences and used to characterise the composition of natural communities developing in young marine biofilms. The possible contribution of the proposed technique in revealing ecological dynamics affecting higher bacterial taxa is discussed., (Copyright © 2011 Elsevier B.V. All rights reserved.)

In order to improve the genetic characterisation of the barnacle Balanus amphitrite, normalised EST libraries for the developmental stages, viz. nauplius (a mix of instars I and II), cyprid and adult, were generated. The libraries were sequenced independently using 454 technologies and 575,666 reads were generated. For adults, 4843 unique isotigs were estimated and 6754 and 7506 in the cyprid and naupliar stage, respectively. It was found that some of the previously proposed cyprid-specific bcs genes were also expressed during the naupliar and adult stage. Furthermore, as lectins have been hypothesised to influence settlement cue recognition in barnacles, the database was searched for lectin-like isotigs. Two proteins, uniquely expressed in either the cyprid or the adult stage, matched a mannose receptor, and their nucleotide sequences were 33% and 31% identical to a lectin (BRA-3) isolated from Megabalanus rosa. Further characterisation of these genes may suggest their involvement in settlement.

Microbial biofilms are composed of a hydrated matrix of biopolymers including polypeptides, polysaccharides and nucleic acids and act as a protective barrier and microenvironment for the inhabiting microbes. While studying marine biofilms, we observed that supernatant produced by a marine isolate of Bacillus licheniformis was capable of dispersing bacterial biofilms. We investigated the source of this activity and identified the active compound as an extracellular DNase (NucB). We have shown that this enzyme rapidly breaks up the biofilms of both Gram-positive and Gram-negative bacteria. We demonstrate that bacteria can use secreted nucleases as an elegant strategy to disperse established biofilms and to prevent de novo formation of biofilms of competitors. DNA therefore plays an important dynamic role as a reversible structural adhesin within the biofilm.

Local field potential (LFP) and Electromyographic (EMG) signals were recorded from 12 Parkinsonian patients with tremor-dominant symptoms as they performed passive and voluntary movements. The LFP signals were categorised into episodes of tremorous and atremorous activity (identified through EMG power spectra), then divided into delta (2-4 Hz), theta (4-8 Hz), alpha (8-13 Hz), and beta (13-30 Hz) frequency bands. Modulation of LFP oscillatory activity in these frequency bands were compared between the subthalamic nucleus (STN) and the globus pallidus internus (GPi) to determine if differential tremor-related characteristics were identifiable for either target. Our results suggest that such local characteristic activity is identifiable in the STN, and thus could be a target for initial development of a closed-loop demand driven stimulator device which capitalises on such activity to trigger stimulation, even during voluntary movement activity., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Sensing their environment is a crucial ability of all life forms. In higher eukaryotes the sensing of airborne volatile compounds, or olfaction, is well developed. In plants, slime moulds and yeast there is also compelling evidence that these organisms can smell their environment and respond accordingly. Here we show that bacteria are also capable of olfaction. Bacillus licheniformis was able to sense airborne volatile metabolites produced by neighbouring bacterial cultures and cells could respond to this chemical information in a coordinated way. When Bacillus licheniformis was grown in a microtitre plate adjacent to a bacterial culture of the same or a different species, growing in complex medium, biofilm formation and pigment production were elicited by volatile molecules. A weaker response occurred in increasingly distant wells. The emitted volatile molecule was identified as ammonia. These data demonstrate that B. licheniformis has evolved the ability collect information about its environment from the surrounding air and physiologically respond to it in a manner similar to olfaction. This is the first time that a behavioural response triggered by odorant molecules received through the gas phase is described in bacteria.

Deep Brain Stimulation (DBS) has been successfully used throughout the world for the treatment of Parkinson's disease symptoms. To control abnormal spontaneous electrical activity in target brain areas DBS utilizes a continuous stimulation signal. This continuous power draw means that its implanted battery power source needs to be replaced every 18-24 months. To prolong the life span of the battery, a technique to accurately recognize and predict the onset of the Parkinson's disease tremors in human subjects and thus implement an on-demand stimulator is discussed here. The approach is to use a radial basis function neural network (RBFNN) based on particle swarm optimization (PSO) and principal component analysis (PCA) with Local Field Potential (LFP) data recorded via the stimulation electrodes to predict activity related to tremor onset. To test this approach, LFPs from the subthalamic nucleus (STN) obtained through deep brain electrodes implanted in a Parkinson patient are used to train the network. To validate the network's performance, electromyographic (EMG) signals from the patient's forearm are recorded in parallel with the LFPs to accurately determine occurrences of tremor, and these are compared to the performance of the network. It has been found that detection accuracies of up to 89% are possible. Performance comparisons have also been made between a conventional RBFNN and an RBFNN based on PSO which show a marginal decrease in performance but with notable reduction in computational overhead.

Bacteria can switch between planktonic forms (single cells) and biofilms, i.e., bacterial communities growing on solid surfaces and embedded in a matrix of extracellular polymeric substance. Biofilm formation by pathogenic bacteria often results in lower susceptibility to antibiotic treatments and in the development of chronic infections; thus, biofilm formation can be considered an important virulence factor. In recent years, much attention has been directed towards understanding the biology of biofilms and towards searching for inhibitors of biofilm development and of biofilm-related cellular processes. In this report, we review selected examples of target-based screening for anti-biofilm agents: We focus on inhibitors of quorum sensing, possibly the most characterized target for molecules with anti-biofilm activity, and on compounds interfering with the metabolism of the signal molecule cyclic di-GMP metabolism and on inhibitors of DNA and nucleotide biosynthesis, which represent a novel and promising class of biofilm inhibitors. Finally, we discuss the activation of biofilm dispersal as a novel mode of action for anti-biofilm compounds.

Domesticated laboratory strains of Bacillus subtilis readily take up and integrate exogenous DNA. In contrast, "wild" ancestors or Bacillus strains recently isolated from the environment can only be genetically modified by phage transduction, electroporation or protoplast transformation. Such methods are laborious, have a variable yield or cannot efficiently be used to alter chromosomal DNA. A major disadvantage of using laboratory strains is that they have often lost, or do not display ecologically relevant physiologies such as the ability to form biofilms. Here we present a method that allows genetic transformation by natural competence in several environmental isolates of B. subtilis. Competence in these strains was established by expressing the B. subtilis competence transcription factor ComK from an IPTG-inducible promoter construct present on an unstable plasmid. This transiently activates expression of the genes required for DNA uptake and recombination in the host strain. After transformation, the comK encoding plasmid is lost easily because of its intrinsic instability and the transformed strain returns to its wild state. Using this method, we have successfully generated mutants and introduced foreign DNA into a number of environmental isolates and also B. subtilis strain NCIB3610, which is widely used to study biofilm formation. Application of the same method to strains of B. licheniformis was unsuccessful. The efficient and rapid approach described here may facilitate genetic studies in a wider array of environmental B. subtilis strains.

Background: Balanus amphitrite is a barnacle commonly used in biofouling research. Although many aspects of its biology have been elucidated, the lack of genetic information is impeding a molecular understanding of its life cycle. As part of a wider multidisciplinary approach to reveal the biogenic cues influencing barnacle settlement and metamorphosis, we have sequenced and annotated the first cDNA library for B. amphitrite. We also present a systematic validation of potential reference genes for normalization of quantitative real-time PCR (qRT-PCR) data obtained from different developmental stages of this animal., Results: We generated a cDNA library containing expressed sequence tags (ESTs) from adult B. amphitrite. A total of 609 unique sequences (comprising 79 assembled clusters and 530 singlets) were derived from 905 reliable unidirectionally sequenced ESTs. Bioinformatics tools such as BLAST, HMMer and InterPro were employed to allow functional annotation of the ESTs. Based on these analyses, we selected 11 genes to study their ability to normalize qRT-PCR data. Total RNA extracted from 7 developmental stages was reverse transcribed and the expression stability of the selected genes was compared using geNorm, BestKeeper and NormFinder. These software programs produced highly comparable results, with the most stable gene being mt-cyb, while tuba, tubb and cp1 were clearly unsuitable for data normalization., Conclusion: The collection of B. amphitrite ESTs and their annotation has been made publically available representing an important resource for both basic and applied research on this species. We developed a qRT-PCR assay to determine the most reliable reference genes. Transcripts encoding cytochrome b and NADH dehydrogenase subunit 1 were expressed most stably, although other genes also performed well and could prove useful to normalize gene expression studies.

Of 131 bacterial isolates from seaweed, a culture of Bacillus licheniformis produced a novel protein with antibacterial activity against methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci, and Listeria monocytogenes. The antibacterial activity was maximal in cultures prepared in Columbia broth containing pieces of synthetic polyurethane sponge and shaken at 210 to 230 rpm. Antibacterial activity was not found in cultures grown statically or with different speeds of rotary shaking. Reduced activity was apparent in supernatants prepared from marine 2216E broth and tryptone soya broth with or without 1% (wt/vol) sodium chloride. The antibacterial compound was sensitive to proteinase K, pronase, and trypsin, but was not affected by Tween-20, -40, -60, or -80, or alpha- or beta-amylase. Activity was not adversely affected by heating up to 40 degrees C or treatment at pH 5 to 14. The bioactive compound was determined to be associated with a protein of 30.7 kDa, which had homology to the YbdN protein of B. licheniformis ATCC 14580.

An endolithic bacterium, strain RSBr-1, was isolated from the inside of a piece of red sandstone from coastal areas of Scotland. RSBr-1 was gram negative, oxidase and catalase positive, and cells were non-motile rods. Sodium was required for growth. The optimum sodium chloride concentration and pH for growth were 4% and pH 8.0, respectively. Eumelanin was produced in marine broth and in BY medium. RSBr-1 hydrolyzes chitin, esculin, gelatin, and starch, but not agar. Nitrate reduction is positive. Taxonomic characterization of this strain indicated that it belongs to the genus Microbulbifer. The difference between the aligned 16S rDNA sequences of RSBr-1 and the closest relative, M. elongata, is greater than the difference between the 16S rDNA sequences of M. hydrolyticus and M. elongata. On the basis of the phenotypic and genotypic comparison of this isolate with the other strains, RSBr-1 is proposed as a new species, Microbulbifer arenaceous, with type strain RSBr-1.

An air-membrane surface (AMS) bioreactor was designed to allow bacteria to grow attached to a surface as a biofilm in contact with air. When Bacillus licheniformis strain EI-34-6, isolated from the surface of a marine alga, was grown in this reactor, cells produced antimicrobial compounds which they did not produce when they were grown in shake flask cultures. An unidentified red pigment was also produced by surface-grown cells but not by planktonically grown cells. Glycerol and ferric iron were important for the production of antimicrobial compounds and the red pigment. Release of these secondary metabolites was not due to the onset of sporulation. Cell-free spent medium recovered from beneath the reactor membrane could induce production of antimicrobial compounds and red pigment in shake flask cultures. Neither glycerol nor ferric iron was required for production of these inducer compounds. Spent medium from beneath the membrane of an AMS bioreactor culture of Bacillus subtilis strain DSM10(T) and Bacillus pumilus strain EI-25-8 could also induce production of antimicrobial compounds and a red pigment in B. licheniformis isolate EI-34-6 grown in shake flask cultures; however, the corresponding spent medium from shake flask cultures of DSM10(T) and EI-25-8 could not. These results suggest that there is a biofilm-specific cross-species signaling system which can induce planktonically grown cells to behave as if they were in a biofilm by regulating the expression of pigments and antimicrobial compounds.

Objectives: This study used a qualitative approach in which participants were asked to write about their experiences in recruiting older women into either one of two exercise intervention studies that are part of the Baltimore Hip Studies. The sample included 8 researcher nurses all women, White, and 42-53 years of age., Background: Older adults, particularly older women, are less likely to participate in research studies when compared to their younger counterparts. The purpose of this study was to explore the techniques successfully used by research nurses in the Baltimore Hip Studies to recruit older women after hip fracture into exercise intervention studies., Method: Data analysis was performed using basic content analysis (Crabtree & Miller, 1992; Miles & Huberman, 1984) "in vivo" coding (Dowd, 1991), or "grounded" coding (Glaser & Strauss, 1967), which involves using the informants' own words to capture a particular idea., Results: A total of 16 codes were identified and reduced to nine themes. Seven themes focused on techniques that facilitated recruitment: (a) caring for individuals; (b) emphasizing benefits; (c) eliciting support from others; (d) being an expert; (e) using role models; (f) using good timing; and (g) giving good first impressions. The remaining two themes identified barriers to recruitment: (a) time commitment and (b) lack of support., Discussion: Based on these themes, specific recruitment techniques are recommended. Ongoing research, however, is needed to establish the most effective recruitment procedures with older women.

Problems with tin and copper antifouling compounds have highlighted the need to develop new environmentally friendly antifouling coatings. Bacteria isolated from living surfaces in the marine environment are a promising source of natural antifouling compounds. Four isolates were used to produce extracts that were formulated into ten water-based paints. All but one of the paints showed activity against a test panel of fouling bacteria. Five of the paints were further tested for their ability to inhibit the settlement of barnacle larvae, Balanus amphitrite, and algal spores of Ulva lactuca, and for their ability to inhibit the growth of U. lactuca. Two paints caused a significant decrease in the number of settled barnacles. One paint containing extract of Pseudomonas sp. strain NUDMB50-11, showed excellent activity in all assays. The antifouling chemicals responsible for the activity of the extract were isolated, using bioassay guided fractionation, and their chemical structures determined.

The bacterial consortium MPD-M, isolated from sediment associated with Colombian mangrove roots, was effective in the treatment of hydrocarbons in water with salinities varying from 0 to 180 g L(-1). Where the salinity of the culture medium surpassed 20 g L(-1), its effectiveness increased when the cells were immobilized on polypropylene fibers. Over the range of salinity evaluated, the immobilized cells significantly enhanced the biodegradation rate of crude oil compared with free-living cells, especially with increasing salinity in the culture medium. Contrary to that observed in free cell systems, the bacterial consortium MPD-M was highly stable in immobilized systems and it was not greatly affected by increments in salinity. Biodegradation was evident even at the highest salinity evaluated (180 g L(-1)), where biodegradation was between 4 and 7 times higher with immobilized cells compared to free cells. The biodegradation of pristane (PR) and phytane (PH) and of the aromatic fraction was also increased using cells immobilized on polypropylene fibers., (Copyright 2002 Wiley Periodicals, Inc.)

Insects produce several types of peptides to combat a broad spectrum of invasive pathogenic microbes, including protozoans. However, despite this defense response, infections are often established. Our aim was to design novel peptides that produce high rates of mortality among protozoa of the genus Plasmodium, the malaria parasites. Using existing antimicrobial peptide sequences as templates, we designed and synthesized three short novel hybrids, designated Vida1 to Vida3. Each has a slightly different predicted secondary structure. The peptides were tested against sporogonic stages of the rodent malaria parasites Plasmodium berghei (in vitro and in vivo) and P. yoelii nigeriensis (in vitro). The level of activity varied for each peptide and according to the parasite stage targeted. Vida3 (which is predicted to have large numbers of beta sheets and coils but no alpha helices) showed the highest level of activity, killing the early sporogonic stages in culture and causing highly significant reductions in the prevalence and intensity of infection of P. berghei after oral administration or injection in Anopheles gambiae mosquitoes. The secondary structures of these peptides may play a crucial role in their ability to interact with and kill sporogonic forms of the malaria parasite.

The known metabolite, enniatin B, and a cyclic tetrapeptide, JM47, which is a new natural product, were extracted from brown rice cultures of a marine fungus, identified as a Fusarium species, isolated from the marine alga Codium fragile. NMR studies, including 15N HMQC and 15N HMBC, established the structure of JM47 as cyclo(Ala-Ala-Aoh-Pro), where Aoh is the amino acid, (2S,9S)-2-amino-8-oxo-9-hydroxydecanoic acid. The absolute stereochemistry of the Aoh side chain carbinol centre was determined using Mosher ester methodology. Analysis of NOESY data assisted by molecular modelling revealed an alternating L-, D-, L-, D-configuration for the tetrapeptide core. The absolute stereochemistry of the core was determined by acidic hydrolysis and chiral TLC analysis of the proline residue. JM47 belongs to the HC-toxin family of cyclic tetrapeptides which possess a 2-amino-8-oxo-9,10-epoxydecanoic acid residue in place of the Aoh unit. This is the first report of an analogue of HC-toxin from a marine Fusarium species.