Extraction of the same novel homoglycan mixture from two different strains of Bifidobacterium animalis and three strains of Bifidobacterium breve.

Three strains of Bifidobacterium breve (JCM 7017, JCM 7019 and JCM 2258) and two strains of Bifidobacterium animalis subsp. lactis (AD011 and A1dOxR) were grown in broth cultures or on plates, and a standard exopolysaccharide extraction method was used in an attempt to recover exocellular polysaccharides. When the extracted materials were analysed by NMR it was clear that mixtures of polysaccharides were being isolated including exopolysaccharides (EPS) cell wall polysaccharides and intracellular polysaccharides. Treatment of the cell biomass from the B. breve strains, or the B. animalis subsp. lactis AD011 strain, with aqueous sodium hydroxide provided a very similar mixture of polysaccharides but without the EPS. The different polysaccharides were partially fractionated by selective precipitation from an aqueous solution upon the addition of increasing percentages of ethanol. The polysaccharides extracted from B. breve JCM 7017 grown in HBM media supplemented with glucose (or isotopically labelled D-glucose-1-13C) were characterised using 1D and 2D-NMR spectroscopy. Addition of one volume of ethanol generated a medium molecular weight glycogen (Mw=1×105 Da, yield 200 mg/l). The addition of two volumes of ethanol precipitated an intimate mixture of a low molecular weight ß-(1→6)-glucan and a low molecular weight ß-(1→6)-galactofuranan which could not be separated (combined yield 46 mg/l). When labelled D-glucose-1-13C was used as a carbon supplement, the label was incorporated into >95% of the anomeric carbons of each polysaccharide confirming they were being synthesised in situ. Similar 1H NMR profiles were obtained for polysaccharides recovered from the cells of B. animalis subsp. lactis AD011and A1dOxR (in combination with an EPS), B. breve JCM 7017, B. breve JCM 7019, B. breve JCM 2258 and from an EPS (-ve) mutant of B. breve 7017 (a non-EPS producer).

Disinfection of artificially contaminated gloved hands reduces transmission of Staphylococcus epidermidis to catheter valves.

Disinfection of gloved hands is advocated increasingly in situations where visibly unsoiled gloves are used during multiple clinical activities on the same patient. As there are no data demonstrating that such practice attributes to lower bacterial transfer during clinical care, a standardized experimental study was conducted. Gloved hands touched chicken breasts contaminated with Staphylococcus epidermidis with or without disinfection before touching sterile catheter valves. Contaminated gloves transferred 5.18 log10 colony-forming units (cfu) S. epidermidis to the catheter valves. Disinfection of contaminated gloves significantly reduced the numbers transferred to 0.78 log10 cfu. Disinfection of gloved hands may reduce the risk of transmission.

The Impact of Alkaliphilic Biofilm Formation on the Release and Retention of Carbon Isotopes from Nuclear Reactor Graphite.

14C is an important consideration within safety assessments for proposed geological disposal facilities for radioactive wastes, since it is capable of re-entering the biosphere through the generation of 14C bearing gases. The irradiation of graphite moderators in the UK gas-cooled nuclear power stations has led to the generation of a significant volume of 14C-containing intermediate level wastes. Some of this 14C is present as a carbonaceous deposit on channel wall surfaces. Within this study, the potential of biofilm growth upon irradiated and 13C doped graphite at alkaline pH was investigated. Complex biofilms were established on both active and simulant samples. High throughput sequencing showed the biofilms to be dominated by Alcaligenes sp at pH 9.5 and Dietzia sp at pH 11.0. Surface characterisation revealed that the biofilms were limited to growth upon the graphite surface with no penetration of the deeper porosity. Biofilm formation resulted in the generation of a low porosity surface layer without the removal or modification of the surface deposits or the release of the associated 14C/13C. Our results indicated that biofilm formation upon irradiated graphite is likely to occur at the pH values studied, without any additional release of the associated 14C.

Floc Formation Reduces the pH Stress Experienced by Microorganisms Living in Alkaline Environments.

The survival of microorganisms within a cementitious geological disposal facility for radioactive wastes heavily depends on their ability to survive the calcium-dominated, hyperalkaline conditions resulting from the dissolution of the cementitious materials. The results from this study show that the formation of flocs, composed of a complex mixture of extracellular polymeric substances (EPS), provides protection against alkaline pH values up to 13.0. The flocs were dominated by Alishewanella and Dietzia spp., producing a mannose-rich carbohydrate fraction incorporating extracellular DNA, resulting in Ca2+ sequestration. EPS provided a ∼10-µm thick layer around the cells within the center of the flocs, which were capable of growth at pH values of 11.0 and 11.5, maintaining internal pH values of 10.4 and 10.7, respectively. Microorganisms survived at a pH of 12.0, where an internal floc pH of 11.6 was observed, as was a reduced associated biomass. We observed limited floc survival (<2 weeks) at a pH of 13.0. This study demonstrates that flocs maintain lower internal pHs in response to the hyperalkaline conditions expected to occur within a cementitious geological disposal facility for radioactive wastes and indicates that floc communities within such a facility can survive at pHs up to 12.0.IMPORTANCE The role of extracellular polymeric substances (EPS) in the survival of microorganisms in hyperalkaline conditions is poorly understood. Here, we present the taxonomy, morphology, and chemical characteristics of an EPS-based microbial floc, formed by a consortium isolated from an anthropogenic hyperalkaline site. Short-term (<2 weeks) survival of the flocs at a pH of 13 was observed, with indefinite survival observed at a pH of 12.0. Measurements from micro-pH electrodes (10-µm-diameter tip) demonstrated that flocs maintain lower internal pHs in response to hyperalkaline conditions (pH 11.0, 11.5, and 12.0), demonstrating that floc formation and EPS production are survival strategies under hyperalkaline conditions. The results indicate how microbial communities may survive and propagate within the hyperalkaline environment that is expected to prevail in a cementitious geological disposal facility for radioactive wastes; the results are also relevant to the wider extremophile community.

A systematic comparison of antimicrobial wound dressings using a planktonic cell and an immobilized cell model.

AIM: The aim of the study was to evaluate the ability of in vitro planktonic and immobilized cell models for determining the antimicrobial efficacy of common antimicrobial wound dressings. METHODS AND RESULTS: Five strains of Acinetobacter baumannii, Pseudomonas aeruginosa and methicillin resistant Staphylococcus aureus were tested against four antimicrobial wound dressings containing silver, honey or polyhexamethylene biguanide (PHMB), using both a planktonic and immobilized cell model. Across all species and models used, the nanocrystalline silver coated dressing demonstrated the best antimicrobial activity being as good if not better than all the other dressings. The planktonic cell model was less effective at differentiating the dressings on antimicrobial performance as the immobilized cell model indicating that a diffusion barrier had a significant impact on the performance of some dressings. In the presence of the diffusion barrier, antimicrobial impact of the Honey and PHMB dressings was significantly reduced, particularly in the case of A. baumannii. Activity was at least an order of magnitude lower in the immobilized cell model vs the planktonic cell model. CONCLUSIONS: The use of a planktonic cell model within standard tests may overestimate the efficacy of honey and PHMB. The use of an immobilized cell model provides a more demanding test for antimicrobial dressings allowing dressing to dressing and pathogen to pathogen differences to be more clearly quantified. SIGNIFICANCE AND IMPACT OF THE STUDY: The introduction of planktonic and immobilized cell models as part of testing regimens for wound dressings will provide a more thorough understanding of their antimicrobial and anti-biofilm properties.

The enrichment of an alkaliphilic biofilm consortia capable of the anaerobic degradation of isosaccharinic acid from cellulosic materials incubated within an anthropogenic, hyperalkaline environment.

Anthropogenic hyperalkaline sites provide an environment that is analogous to proposed cementitious geological disposal facilities (GDF) for radioactive waste. Under anoxic, alkaline conditions cellulosic wastes will hydrolyze to a range of cellulose degradation products (CDP) dominated by isosaccharinic acids (ISA). In order to investigate the potential for microbial activity in a cementitious GDF, cellulose samples were incubated in the alkaline (∼pH 12), anaerobic zone of a lime kiln waste site. Following retrieval, these samples had undergone partial alkaline hydrolysis and were colonized by a Clostridia-dominated biofilm community, where hydrogenotrophic, alkaliphilic methanogens were also present. When these samples were used to establish an alkaline CDP fed microcosm, the community shifted away from Clostridia, methanogens became undetectable and a flocculate community dominated by Alishewanella sp. established. These flocs were composed of bacteria embedded in polysaccharides and proteins stabilized by extracellular DNA. This community was able to degrade all forms of ISA with >60% of the carbon flow being channelled into extracellular polymeric substance (EPS) production. This study demonstrated that alkaliphilic microbial communities can degrade the CDP associated with some radioactive waste disposal concepts at pH 11. These communities divert significant amounts of degradable carbon to EPS formation, suggesting that EPS has a central role in the protection of these communities from hyperalkaline conditions.

Testing standards for sporicides.

Sporicidal products are of considerable importance in healthcare environments due to the requirement for products that are capable of dealing with contamination with Clostridium difficile spores. Sporicidal testing standards to validate the claims of sporicidal activity are an important tool in the evaluation of commercial sporicides. Within Europe there are a number of sporicidal testing standards which are often used to validate the claims of commercial sporicides. However, the extent to which these standards reflect the practical application of sporicides in healthcare settings is limited since they employ long contact times (≥30min) and do not involve surface contamination. Alternative international standards are available which employ contaminated carriers rather than spore suspensions, and the Organisation for Economic Co-operation and Development is currently developing a unified set of standards which are more realistic in their design than the currently available European standards. This paper reviews the currently available testing standards for sporicides, highlighting the key procedural differences between them and the extent to which they reflect the practical application of sporicidal products. Some of the common problems and errors associated with the application of the European sporicidal standard methods are also highlighted and discussed. Finally gaps in the currently available testing standards are identified and discussed.

Assessing the potential of short rotation coppice (SRC) for cleanup of radionuclide-contaminated sites.

A small-scale greenhouse investigation was undertaken using Goat willow (Salix caprea) and aspen (Populus tremula) to evaluate the potential of short rotation coppice for remediation of 137Cs- and 90Sr-contaminated sites. Results showed that both species were able to accumulate these radionuclides from a representative disposal soil (aged) and a spiked soil S. caprea accumulating greater levels of 137Cs than P. tremula, with no difference between species for 90Sr accumulation. For each radionuclide, the distribution in both species was similar, with 137Cs accumulation greatest in the roots, whereas 90Sr accumulation was greatest in the leaves. It was also evident that the soil-to-plant transfer factor (Tf) values for 90Sr were greater than for 137Cs, agreeing with differences in the reported bioavailailablity of these radionuclides in soil Based on the Tf values for S. caprea (conservative), estimated remediation times were 92 and 56 yr, for 137Cs and 90Sr, respectively. It is suggested that the selection of Salix species grown in a system of SRC provides a significant opportunity for removal of both 137Cs and 90Sr, primarily due to its higher biomass production. However, for 137Cs phytoremediation investigations into the appropriate use of soil amendments for increasing bioavailability are required.

Ultrastructure of the budgerigar testis during a photoperiodically induced cycle.

Spermatogenesis in the Budgerigar can be arrested by reducing the birds' photoperiod to 8 hours of daylight or less. When this occurs, Sertoli cell cytoplasm shows a great increase in the size and number of residual bodies, while the smooth endoplasmic reticulum is reduced. If the bird is kept at 8 hours of daylight for some weeks large lipid droplets are seen in Sertoli cytoplasm, and degenerated spermatids are apparently phagocytosed. The interstitium shows fewer active Leydig cells, a paucity of lipids and occasional ovoid mitochondria. The basal lamellae of the tubule which are thick and convoluted before and during spermatogenesis become thinner and straighter. It is thought that these morphological changes reflect changes in metabolic activity.

The differentiation of the acrosome in the spermatid of the budgerigar (Melopsittacus undulatus).

The development of the acrosome of the budgerigar spermatid was studied under the electronmicroscope. The acrosome arises from a granule derived from the Golgi apparatus which interacts with the nucleus to form a cone shaped acrosomal cap and a perforatorium. A quanity of cytoplasm and plasma membrane is captured from the cell periphery and incorporated into the substance of the acrosome. The possible significance of the developmental phases and comparative avian spermatology is discussed.