Efficacy of Biocides Used in the Modern Food Industry To Control Salmonella enterica, and Links between Biocide Tolerance and Resistance to Clinically Relevant Antimicrobial Compounds

Biocides play an essential role in limiting the spread of infectious disease. The food industry is dependent on these agents, and their increasing use is a matter for concern. Specifically, the emergence of bacteria demonstrating increased tolerance to biocides, coupled with the potential for the development of a phenotype of cross-resistance to clinically important antimicrobial compounds, needs to be assessed. In this study, we investigated the tolerance of a collection of susceptible and multidrug-resistant (MDR) Salmonella enterica strains to a panel of seven commercially available food-grade biocide formulations. We explored their abilities to adapt to these formulations and their active biocidal agents, i.e., triclosan, chlorhexidine, hydrogen peroxide, and benzalkonium chloride, after sequential rounds of in vitro selection. Finally, cross-tolerance of different categories of biocidal formulations, their active agents, and the potential for coselection of resistance to clinically important antibiotics were investigated. Six of seven food-grade biocide formulations were bactericidal at their recommended working concentrations. All showed a reduced activity against both surface-dried and biofilm cultures. A stable phenotype of tolerance to biocide formulations could not be selected. Upon exposure of Salmonella strains to an active biocidal compound, a high-level of tolerance was selected for a number of Salmonella serotypes. No cross-tolerance to the different biocidal agents or food-grade biocide formulations was observed. Most tolerant isolates displayed changes in their patterns of susceptibility to antimicrobial compounds. Food industry biocides are effective against planktonic Salmonella. When exposed to sublethal concentrations of individual active biocidal agents, tolerant isolates may emerge. This emergence was associated with changes in antimicrobial susceptibilities.

A morphologic and autoradiographic study of cell death and regeneration in the retinal microvasculature of normal and diabetic rats

Cell loss and regeneration were investigated and compared in the retinal microvasculature of age- and sex-matched normal and streptozotocin diabetic rats. Selective pericyte loss in the diabetic rat was characterized by changes in the pericyte to endothelial cell ratio in retinal capillaries isolated for microscopy by the trypsin digest technique. A comparison of 3- and 9-month-old normal rats showed no significant change in the pericyte to endothelial cell ratio (1:2.7). In diabetic animals the ratio was reduced to 1:4.03, which was statistically significant (P less than .001). Premitotic retinal vascular cells in normal and diabetic rats were labelled with tritiated thymidine and the labelling indices calculated from cell counts of trypsin digest preparations. Methyl H3 thymidine was infused continuously over an eight-day period using osmotic mini pumps. The labelling index of endothelial cells (0.33%) in normal rats increased to 0.91% in diabetic animals (P less than .05). The labelling index of pericyte cells in normal animals (0.16%) did not increase significantly (P greater than .05) in diabetic animals (0.19%). A special stain was used to exclude labelled polymorphonuclear leukocytes from the cell counts.

Increased susceptibility of a triclabendazole (TCBZ)-resistant isolate of Fasciola hepatica to TCBZ following co-incubation in vitro with the P-glycoprotein inhibitor, R(+)-verapamil

SUMMARY A study was carried out to investigate whether the action of triclabendazole sulphoxide (TCBZ.SO) against the liver fluke, Fasciola hepatica is altered by inhibition of P-glycoprotein (Pgp)-linked drug efflux pumps. The Oberon TCBZ-resistant and Cullompton TCBZ-susceptible fluke isolates were used for this in vitro study and the Pgp inhibitor selected was R(+)-verapamil [R(+)-VPL]. For experiments with the Oberon isolate, flukes were incubated for 24 h with either R(+)-VPL (1×10-4 m) on its own, TCBZ.SO (15 µg mL-1) alone, a combination of R(+)-VPL (1×10-4 m) plus TCBZ.SO (15 µg mL-1), TCBZ.SO (50 µg mL-1) on its own, or a combination of TCBZ.SO (50 µg mL-1) plus R(+)-VPL (1×10-4 m). They were also incubated in TCBZ.SO (50 µg mL-1) alone or in combination with R(+)-VPL (1×10-4 m) until they became inactive; and in TCBZ.SO (50 µg mL-1) alone for a time to match that of the combination inactivity time. Flukes from the Cullompton isolate were treated with either TCBZ.SO (50 µg mL-1) alone or in combination with R(+)-VPL (1×10-4 m) until they became inactive, or with TCBZ.SO (50 µg mL-1) alone time-matched to the combination inactivity time. Morphological changes resulting from drug treatment and following Pgp inhibition were assessed by means of scanning electron microscopy. Incubation in R(+)-VPL alone had a minimal effect on either isolate. TCBZ.SO treatment had a relatively greater impact on the TCBZ-susceptible Cullompton isolate. When R(+)-VPL was combined with TCBZ.SO in the incubation medium, however, the surface disruption to both isolates was more severe than that seen after TCBZ.SO treatment alone; also, the time taken to reach inactivity was shorter. More significantly, though, the potentiation of drug activity was greater in the Oberon isolate; also, it was more distinct at the higher concentration of TCBZ.SO. So, the Oberon isolate appears to be particularly sensitive to efflux pump inhibition. The results of this study suggest that enhanced drug efflux in the Oberon isolate may be involved in the mechanism of resistance to TCBZ.

Effect of the P-glycoprotein inhibitor, R(+)-verapamil on the drug susceptibility of a triclabendazole-resistant isolate of Fasciola hepatica

A study has been carried out to investigate whether the action of triclabendazole (TCBZ) against Fasciola hepatica is altered by the inhibition of P-glycoprotein (Pgp)-linked drug efflux pumps. The Sligo TCBZ-resistant and Cullompton TCBZ-susceptible fluke isolates were used for these experiments and the Pgp inhibitor selected was R(+)-verapamil [R-VPL]. In the first experiment, flukes were initially incubated for 2 h in R-VPL (100 µM), then incubated for a further 22 h in R-VPL+triclabendazole sulphoxide (TCBZ.SO) (50 µg/ml, or 0.1327 µM). For controls, flukes were incubated for 24 h in R-VPL and TCBZ.SO on their own. In a second experiment, flukes were removed from the incubation media following cessation of movement. In the third experiment, Sligo flukes were incubated in lower concentrations of R-VPL (10 µM) and TCBZ.SO (15 µg/ml, or 0.0398 µM). Morphological changes resulting from drug treatment and following Pgp inhibition were assessed by means of scanning electron microscopy. Incubation in R-VPL alone had minimal effect on either isolate. After treatment with TCBZ.SO alone, there was greater surface disruption to the Cullompton than Sligo isolate. However, combined treatment of R-VPL+TCBZ.SO led to more severe surface changes to the Sligo isolate than with TCBZ.SO on its own; this potentiation of drug activity was not seen with the Cullompton isolate. The phenomenon was evident at both concentrations of TCBZ.SO. Inclusion of R-VPL in the incubation medium also reduced the time taken for the flukes to become inactive; again, this effect was more distinct with the Sligo isolate. The results of this study support the concept of altered drug efflux in TCBZ-resistant flukes and indicate that drug transporters may play a role in the development of drug resistance.

Impact of Heat Pump Load on Distribution Networks

Heat pumps can provide domestic heating at a cost that is competitive with oil heating in particular. If the electricity supply contains a significant amount of renewable generation, a move from fossil fuel heating to heat pumps can reduce greenhouse gas emissions. The inherent thermal storage of heat pump installations can also provide the electricity supplier with valuable flexibility. The increase in heat pump installations in the UK and Europe in the last few years poses a challenge for low-voltage networks, due to the use of induction motors to drive the pump compressors. The induction motor load tends to depress voltage, especially on starting. The paper includes experimental results, dynamic load modelling, comparison of experimental results and simulation results for various levels of heat pump deployment. The simulations are based on a generic test network designed to capture the main characteristics of UK distribution system practice. The simulations employ DIgSlILENT to facilitate dynamic simulations that focus on starting current, voltage variations, active power, reactive power and switching transients.

Halophilic microorganisms as a source of novel antibiofilm compounds

Objectives: There is great urgency for alternate sources of antibiotics to be identified. One relatively untapped source of novel bioproducts, including antimicrobials, is organisms derived from extreme environments. Halophiles (which require high salt concentrations) are one such group which is being increasingly explored for their biotechnological potential. The aim of this study was to identify halophilic environmental isolates which possessed in vitro and in vivo antimicrobial and antibiofilm activities. Methods: 73 halophilic bacteria and archaea were isolated from Kilroot salt mine in Northern Ireland. Culture extracts of each isolate were screened for antimicrobial and antibiofilm activity against numerous pathogenic bacteria, including Staphylococcus species and Pseudomonas aeruginosa, both model strains and clinical isolates. The methods used included disc diffusion assays of crude extracts, MIC screening, the MBEC assay, and an in vivo model based on the Greater Wax Moth (Galleria mellonella). Results: The assays indicated >50% of extracts displayed antimicrobial and antibiofilm activity against at least one pathogen, the majority being Staphylococcus species, but also E. coli and P. aeruginosa. Biofilms were either reduced or eradicated by halophile extracts when tested with the MBEC device. Further experiments demonstrated that these effects could be replicated in vivo, with extracts reducing the severity of infections and enhancing the survival of infected G. mellonella. Conclusions: The importance of extremophiles to pharmaceutical research should not be underestimated. While not yet fully characterised, based on the data obtained, the halophiles isolated during this study may provide a promising reservoir of novel antimicrobial and antibiofilm compounds.

Autoecological Approaches to Resolve Subjective Taxonomic Divisions within Arcellacea

Arcellacea (testate lobose amoebae) are important lacustrine environmental indicators that have been used in paleoclimatic reconstructions, assessing the effectiveness of mine tailings pond reclamation projects and for studying the effects of land use change in rural, industrial and urban settings. Recognition of ecophenotypically significant infra-specific ‘strains’ within arcellacean assemblages has the potential to enhance the utility of the group in characterizing contemporary and paleoenvironments. We present a novel approach which employs statistical tools to investigate the environmental and taxonomic significance of proposed strains. We test this approach on two identified strains: Difflugia protaeiformis Lamarck strain ‘acuminata’ (DPA), characterized by fine grained agglutination, and Difflugia protaeiformis Lamarck strain ‘claviformis’ (DPC), characterized by coarse grained agglutination. Redundancy analysis indicated that both organisms are associated with similar environmental variables. No relationship was observed between substrate particle size and abundance of DPC, indicating that DPC has a size preference for xenosomes during test construction. Thus DPC should not be designated as a distinct strain but rather form a species complex with DPA. This study elucidates the need to justify the designation of strains based on their autecology in addition to morphological stability.

A single-component multidrug transporter of the major facilitator superfamily is part of a network that protects Escherichia coli from bile salt stress

Resistance to high concentrations of bile salts in the human intestinal tract is vital for the survival of enteric bacteria such as Escherichia coli. Although the tripartite AcrAB-TolC efflux system plays a significant role in this resistance, it is purported that other efflux pumps must also be involved. We provide evidence from a comprehensive suite of experiments performed at two different pH values (7.2 and 6.0) that reflect pH conditions that E. coli may encounter in human gut that MdtM, a single-component multidrug resistance transporter of the major facilitator superfamily, functions in bile salt resistance in E. coli by catalysing secondary active transport of bile salts out of the cell cytoplasm. Furthermore, assays performed on a chromosomal ΔacrB mutant transformed with multicopy plasmid encoding MdtM suggested a functional synergism between the single-component MdtM transporter and the tripartite AcrAB-TolC system that results in a multiplicative effect on resistance. Substrate binding experiments performed on purified MdtM demonstrated that the transporter binds to cholate and deoxycholate with micromolar affinity, and transport assays performed on inverted vesicles confirmed the capacity of MdtM to catalyse electrogenic bile salt/H(+) antiport.

Properties of two interlinked Chi(2) interactions in noncollinear phase matching

We present a compact experimental realization of the interaction among five field modes in a chi((2)) nonlinear crystal. The classical evolution of the fields can be analytically described assuming that two of the fields play the role of nondepleted pumps. A peculiar behavior appears that has been experimentally verified. If one of the fields has a nonzero input amplitude, then the other two fields at the output are holographic replicas of the input signal. (C) 2004 Optical Society of America

Energy monitoring and quality control of a single screw extruder

Polymer extrusion, in which a polymer is melted and conveyed to a mould or die, forms the basis of most polymer processing techniques. Extruders frequently run at non-optimised conditions and can account for 15–20% of overall process energy losses. In times of increasing energy efficiency such losses are a major concern for the industry. Product quality, which depends on the homogeneity and stability of the melt flow which in turn depends on melt temperature and screw speed, is also an issue of concern of processors. Gear pumps can be used to improve the stability of the production line, but the cost is usually high. Likewise it is possible to introduce energy meters but they also add to the capital cost of the machine. Advanced control incorporating soft sensing capabilities offers opportunities to this industry to improve both quality and energy efficiency. Due to strong correlations between the critical variables, such as the melt temperature and melt pressure, traditional decentralized PID (Proportional–Integral–Derivative) control is incapable of handling such processes if stricter product specifications are imposed or the material is changed from one batch to another. In this paper, new real-time energy monitoring methods have been introduced without the need to install power meters or develop data-driven models. The effects of process settings on energy efficiency and melt quality are then studied based on developed monitoring methods. Process variables include barrel heating temperature, water cooling temperature, and screw speed. Finally, a fuzzy logic controller is developed for a single screw extruder to achieve high melt quality. The resultant performance of the developed controller has shown it to be a satisfactory alternative to the expensive gear pump. Energy efficiency of the extruder can further be achieved by optimising the temperature settings. Experimental results from open-loop control and fuzzy control on a Killion 25 mm single screw extruder are presented to confirm the efficacy of the proposed approach.

Disruption of vitellogenesis and spermatogenesis by triclabendazole (TCBZ) in a TCBZ-resistant isolate of Fasciola hepatica following incubation in vitro with a P-glycoprotein inhibitor

A study has been carried out to investigate whether the action of triclabendazole (TCBZ) against Fasciola hepatica is altered by inhibition of P-glycoprotein (Pgp)-linked drug efflux pumps. The Sligo TCBZ-resistant fluke isolate was used for these experiments and the Pgp inhibitor selected was R(+)-verapamil [R(+)-VPL]. In the first experiment, flukes were initially incubated for 2 h in R(+)-VPL (100 μ m), then incubated in R(+)-VPL+triclabendazole sulphoxide (TCBZ.SO) (50 μg mL-1, or 133·1 μ m) until flukes ceased movement (at 9 h post-treatment). In a second experiment, flukes were incubated in TCBZ.SO alone and removed from the incubation medium following cessation of motility (after 15 h). In the third experiment, flukes were incubated for 24 h in R(+)-VPL on its own. Changes to the testis tubules and vitelline follicles following drug treatment and following Pgp inhibition were assessed by means of light microscope histology and transmission electron microscopy. Incubation of the Sligo isolate in either R(+)-VPL or TCBZ.SO on their own had a limited impact on the morphology of the two tissues. Greater disruption was observed when the drugs were combined, in terms of the block in development of the spermatogenic and vitelline cells and the apoptotic breakdown of the remaining cells. Sperm formation was severely affected and abnormal. Large spaces appeared in the vitelline follicles and synthesis of shell protein was disrupted. The results of this study support the concept of altered drug efflux in TCBZ-resistant flukes and indicate that drug transporters may play a role in the development of drug resistance.

Microbial weeds in hypersaline habitats: the enigma of the weed-like Haloferax mediterranei

Heterotrophic prokaryotic communities that inhabit saltern crystallizer ponds are typically dominated by two species, the archaeon Haloquadratum walsbyi and the bacterium Salinibacter ruber, regardless of location. These organisms behave as ‘microbial weeds’ as defined by Cray et al. (Microb Biotechnol 6: 453–492, 2013) that possess the biological traits required to dominate the microbiology of these open habitats. Here, we discuss the enigma of the less abundant Haloferax mediterranei, an archaeon that grows faster than any other, comparable extreme halophile. It has a wide window for salt tolerance, can grow on simple as well as on complex substrates and degrade polymeric substances, has different modes of anaerobic growth, can accumulate storage polymers, produces gas vesicles, and excretes halocins capable of killing other Archaea. Therefore, Hfx. mediterranei is apparently more qualified as a ‘microbial weed’ than Haloquadratum and Salinibacter. However, the former differs because it produces carotenoid pigments only in the lower salinity range and lacks energy-generating retinal-based, light-driven ion pumps such as bacteriorhodopsin and halorhodopsin. We discuss these observations in relation to microbial weed biology in, and the open-habitat ecology of, hypersaline systems.

Quantitative Analysis of Fault and Fracture Systems and their Impact on Groundwater Flow in Irish Bedrock Aquifers

Fault and fracture systems are the most important store and pathway for groundwater in Ireland’s bedrock aquifers, either directly as conductive flow structures, or indirectly as the locus for the development of dolomitised limestone and karst. This article presents the preliminary results of a study involving the quantitative analysis of fault and fracture systems in the broad range of Irish bedrock types and a consideration of their impact on groundwater flow. The principal aims of the project are to develop generic conceptual models for different fault/fracture systems in different lithologies and at different depths, and to link them to observed groundwater behaviour. Here we briefly describe the geometrical characteristics of the main post-Devonian fault/fracture systems controlling groundwater flow from field observations at outcrops, quarries and mines. The structures range from Lower Carboniferous normal faults through to Variscan-related faults and veins, with the most recent structures including Tertiary strike-slip faults and ubiquitous uplift-related joint systems. The geometrical characteristics of different fault/fracture systems combined with observations of groundwater behaviour in both quarry and mine localities, can be linked to general flow and transport conceptualisations of Irish fractured bedrock. Most importantly they also provide a basis for relating groundwater flow to particular fault/fracture systems and their expression with depth and within different lithological sequences, as well as their regional variability.

Lead in rice: Analysis of baseline lead levels in market and field collected rice grains

In a large scale survey of rice grains from markets (13 countries) and fields (6 countries), a total of 1578 rice grain samples were analysed for lead. From the market collected samples, only 0.6% of the samples exceeded the Chinese and EU limit of 0.2 μg g− 1 lead in rice (when excluding samples collected from known contaminated/mine impacted regions). When evaluating the rice grain samples against the Food and Drug Administration's (FDA) provisional total tolerable intake (PTTI) values for children and pregnant women, it was found that only people consuming large quantities of rice were at risk of exceeding the PTTI from rice alone. Furthermore, 6 field experiments were conducted to evaluate the proportion of the variation in lead concentration in rice grains due to genetics. A total of 4 of the 6 field experiments had significant differences between genotypes, but when the genotypes common across all six field sites were assessed, only 4% of the variation was explained by genotype, with 9.5% and 11% of the variation explained by the environment and genotype by environment interaction respectively. Further work is needed to identify the sources of lead contamination in rice, with detailed information obtained on the locations and environments where the rice is sampled, so that specific risk assessments can be performed.

Investigation of the process energy demand in polymer extrusion: a brief review and an experimental study

Extrusion is one of the fundamental production methods in the polymer processing industry and is used in the production of a large number of commodities in a diverse industrial sector. Being an energy intensive production method, process energy efficiency is one of the major concerns and the selection of the most energy efficient processing conditions is a key to reducing operating costs. Usually, extruders consume energy through the drive motor, barrel heaters, cooling fans, cooling water pumps, gear pumps, etc. Typically the drive motor is the largest energy consuming device in an extruder while barrel/die heaters are responsible for the second largest energy demand. This study is focused on investigating the total energy demand of an extrusion plant under various processing conditions while identifying ways to optimise the energy efficiency. Initially, a review was carried out on the monitoring and modelling of the energy consumption in polymer extrusion. Also, the power factor, energy demand and losses of a typical extrusion plant were discussed in detail. The mass throughput, total energy consumption and power factor of an extruder were experimentally observed over different processing conditions and the total extruder energy demand was modelled empirically and also using a commercially available extrusion simulation software. The experimental results show that extruder energy demand is heavily coupled between the machine, material and process parameters. The total power predicted by the simulation software exhibits a lagging offset compared with the experimental measurements. Empirical models are in good agreement with the experimental measurements and hence these can be used in studying process energy behaviour in detail and to identify ways to optimise the process energy efficiency.