Trends in the epidemiology of Candida bloodstream infections in Northern Ireland between January 1984 and December 2000

Objective: To describe the epidemiology of Candida bloodstream infections (BSI) in Northern Ireland. Methods: Retrospective collation of data relating to all clinically significant BSI in a university teaching hospital, which had been recorded prospectively, between 1984 and 2000. Results: One hundred and forty five episodes of candidaemia occurred in 144 patients (of mean age 56.6 years). The contribution of Candida spp. towards all significant BSI increased from 2.00% to 2.5%. C. albicans was the most frequently isolated species, however, its incidence fell from 70% to 53% during the study period. The greatest increase in incidence was seen with C. glabrata which was the most common non-albicans species. Twenty-nine per cent of isolates occurred in patients from an intensive care unit and, surprisingly, a further 25.5% occurred in patients from a surgical service. Conclusion: There appears to be several subtle differences in the epidemiology of candidal BSI between Northern Ireland and other countries. © 2002 The British Infection Society.

Trends in candidemia and antifungal susceptibility in a university hospital in Northern Ireland 2001-2006

Objectives: To describe the species distribution and antifungal susceptibility trends for documented episodes of candidemia at the Royal Hospitals, Belfast, 2001-2006. Methods: Laboratory-based retrospective observational study of all episodes of candidemia. Results: There were 151 episodes of candidemia. The species recovered were: 96 C. albicans; 26 C. glabrata; 18 C. parapsilosis; five C. tropicalis; four C. guilliermondii; one C. famata and one C. dubliniensis. We separated the data into two periods 2001-2003 and 2004-2006; contrary to the findings of other investigators, there was a notable trends toward increasing frequency of C. albicans and decreasing frequency of non-albicans species over time. Although the proportion of C. albicans, C. parapsilosis and C. tropicalis isolates susceptible to fluconazole was unchanged over time, a trend of decreased susceptibility of C. glabrata to fluconazole was noted over the six-year period. Overall, 73% and 7.7% of C. glabrata isolates had susceptible-dose-dependent and resistant phenotypes, respectively. The percentage of C. glabrata isolates susceptible to fluconazole (MIC

Catheter-related bloodstream infection in adult haematology patients:Catheter removal practice and outcome

The aim of the present study was to describe the practice of central venous catheter (CVC) removal and outcomes of catheter-related bloodstream infection (CR-BSI) in adult haematology patients. Patients were identified retrospectively according to diagnosis coding of inpatient episodes and evaluated when, on examination of medical records, there had been evidence of sepsis with strong clinical suspicion that the source was the CVC. Demographic and bacteriological data, as well as therapeutic measures and clinical outcomes, were recorded. One hundred and three patient episodes were evaluated. The most frequent type of CVC was the Hickman catheter and the most frequently isolated pathogen was coagulase-negative staphylococci. Twenty-five percent of episodes were managed with catheter removal. Treatment failure, defined as recurrence of infection within 90 days or mortality attributed to sepsis within 30 days, occurred significantly more frequently in the group managed without catheter removal (52.5% versus 4%, P

Genotypic subgrouping of clinical isolates of Candida albicans and Candida dubliniensis by 25S intron analysis

To determine the frequency, distribution and association of genotypes of Candida albicans and C. dubliniensis in invasive and noninvasive clinical isolates.

Flux and turnover of fixed carbon in soil microbial biomass of limed and unlimed plots of an upland grassland ecosystem

The influence of liming on rhizosphere microbial biomass C and incorporation of root exudates was studied in the field by in situ pulse labelling of temperate grassland vegetation with (13)CO(2) for a 3-day period. In plots that had been limed (CaCO(3) amended) annually for 3 years, incorporation into shoots and roots was, respectively, greater and lower than in unlimed plots. Analysis of chloroform-labile C demonstrated lower levels of (13)C incorporation into microbial biomass in limed soils compared to unlimed soils. The turnover of the recently assimilated (13)C compounds was faster in microbial biomass from limed than that from unlimed soils, suggesting that liming increases incorporation by microbial communities of root exudates. An exponential decay model of (13)C in total microbial biomass in limed soils indicated that the half-life of the tracer within this carbon pool was 4.7 days. Results are presented and discussed in relation to the absolute values of (13)C fixed and allocated within the plant-soil system.

Development and characterization of a lux-modified 2,4-dichlorophenol-degrading Burkholderia sp. RASC

lux-marked biosensors for assessing the toxicity and bioremediation potential of polluted environments may complement traditional chemical techniques. luxCDABE genes were introduced into the chromosome of the 2,4-dichlorophenol (2,4-DCP)-mineralizing bacterium, Burkholderia sp. RASC c2, by biparental mating using the Tn4431 system. Experiments revealed that light output was constitutive and related to cell biomass concentration during exponential growth. The transposon insertion was stable and did not interrupt 2,4-DCP-degradative genes, and expression of luxCDABE did not constitute a metabolic burden to the cell. A bioluminescence response was detectable at sublethal 2,4-DCP concentrations: at <10.26 microg ml(-1), bioluminescence was stimulated (e.g. 218% of control), but at concentrations >60 microg ml(-1) it declined to <1%. Investigating the effect of [14C]-2,4-DCP concentration on the evolution of 14CO2 revealed that, for initial concentrations of 2.5-25 microg ml(-1), approximately equals 55% of the added 14C was mineralized after 24 h compared with

Lux-biosensor assessment of pH effects on microbial sorption and toxicity of chlorophenols

Lux-marked bacterial biosensors and a commercial toxicity testing bacterial strain (Microtox) were exposed to 2,4-dichlorophenol (DCP) and the light output response measured. Increasing DCP concentrations caused a decrease in light output in all three biosensors with an order of sensitivity (in terms of luminescence decrease over the DCP concentration range) of Pseudomonas fluorescens <Escherichia coli <Microtox. Adsorption of DCP to E. coli was measured using uniformly ring labelled [14C]DCP and found to be very rapid. The effect of pH on toxicity and adsorption was also investigated. Low pH values increased the amount of DCP adsorbed to the cell and increased the toxicity of DCP.

Production and evaluation of the utility of novel phage display-derived peptide ligands to Salmonella spp. for magnetic separation

Aims: The objectives of this study were to produce Salmonella-specific peptide ligands by phage display biopanning and evaluate their use for magnetic separation (MS).
Methods and Results: Four phage display biopanning rounds were performed and the peptides expressed by the two most Salmonella-specific (on the basis of phage binding ELISA results) phage clones, MSal020401 and MSal020417, were chemically synthesized and coupled to MyOne™ tosylactivated Dynabeads®. Peptide capture capability for whole Salmonella cells from non-enriched broth cultures was quantified by MS + plate counts and MS + Greenlight™ detection, and compared to capture capability of anti-Salmonella (antibody-coated) Dynabeads®. MS + Greenlight™ gave a more comprehensive picture of capture capability than MS + plate counts and showed that Peptide MSal020417-coated beads exhibited at least similar, if not better, capture capability to anti-Salmonella Dynabeads® (mean capture values of 36.0 ± 18.2 % and 31.2 ± 20.1 %, respectively, over Salmonella spp. concentration range 3 x 101 - 3 x 106 cfu ml-1) with minimal cross-reactivity (= 1.9 %) to three other foodborne bacteria.
Conclusions: One of the phage display-derived peptide ligands was demonstrated by MS + Greenlight™ to be a viable antibody-alternative for MS of Salmonella spp.
Significance and Impact of Study: This study demonstrates an antibody-free approach to Salmonella detection and opens substantial possibilities for more rapid tests for this bacterium.

Effects of alcohols and compatible solutes on the activity of β-galactosidase

During alcoholic fermentation, the products build up and can, ultimately, kill the organism due to their effects on the cell's macromolecular systems. The effects of alcohols on the steady-state kinetic parameters of the model enzyme ß-galactosidase were studied. At modest concentrations (0 to 2 M), there was little effect of methanol, ethanol, propanol and butanol on the kinetic constants. However, above these concentrations, each alcohol caused the maximal rate, V (max), to fall and the Michaelis constant, K (m), to rise. Except in the case of methanol, the chaotropicity of the solute, rather than its precise chemical structure, determined and can, therefore, be used to predict inhibitory activity. Compounds which act as compatible solutes (e.g. glycerol and other polyols) generally reduced enzyme activity in the absence of alcohols at the concentration tested (191 mM). In the case of the ethanol- or propanol-inhibited ß-galactosidase, the addition of compatible solutes was unable to restore the enzyme's kinetic parameters to their uninhibited levels; addition of chaotropic solutes such as urea tended to enhance the effects of these alcohols. It is possible that the compatible solutes caused excessive rigidification of the enzyme's structure, whereas the alcohols disrupt the tertiary and quaternary structure of the protein. From the point of view of protecting enzyme activity, it may be unwise to add compatible solutes in the early stages of industrial fermentations; however, there may be benefits as the alcohol concentration increases.

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.

Development and Validation of a PulseNet Standardized Protocol for Subtyping Isolates of Cronobacter Species

Cronobacter (formerly known as Enterobacter sakazakii) is a genus comprising seven species regarded as opportunistic pathogens that can be found in a wide variety of environments and foods, including powdered infant formula (PIF). Cronobacter sakazakii, the major species of this genus, has been epidemiologically linked to cases of bacteremia, meningitis in neonates, and necrotizing enterocolitis, and contaminated PIF has been identified as an important source of infection. Robust and reproducible subtyping methods are required to aid in the detection and investigation, of foodborne outbreaks. In this study, a pulsed-field gel electrophoresis (PFGE) protocol was developed and validated for subtyping Cronobacter species. It was derived from an existing modified PulseNet protocol, wherein XbaI and SpeI were the primary and secondary restriction enzymes used, generating an average of 14.7 and 20.3 bands, respectively. The PFGE method developed was both reproducible and discriminatory for subtyping Cronobacter species.

Effect of pH on the in vitro susceptibility of planktonic and biofilm-grown Proteus mirabilis to the quinolone antimicrobials

To examine the effect of elevated pH, as reported during urinary catheter infections, on quinolone activity against the urease-producing pathogen Proteus mirabilis.

Rapid and efficient production of L-lactate from xylose using electrodialysis culture-associated product separation

L-Lactate was produced from xylose using electrodialysis culture (ED-C)-associated product separation. In a medium containing 50 g xylose/l, the ED-C was completed in only 32 h (i.e. less than half the time taken by the control culture, without electrodialysis). At 80 g xylose/l, the control culture was unable to consume more than 50 g xylose/1, whereas the ED-C showed increased xylose consumption and was completed by 45 h. The maximum rate of lactate production in the ED-C was higher than that in the control culture. ED-C was also carried out (at 80 g initial xylose/ l) with a supply of fresh xylose-free medium. This ED-C was completed within 30 h, which represents a reduction in fermentation time of 15 h when compared to ED-C without addition of xylose-free medium. Thus, rapid production of L-lactate was achieved by using ED-C which supplied fresh xylose-free medium.

Ethanol-induced water stress and fungal growth

Fungal growth inhibition by ethanol was compared with that caused by five other agents of water stress (at 25, 40 and 42.5°C), using Aspergillus oryzae. Ethanol, KCl, glycerol, glucose, sorbitol, and polyethylene glycol 400 were incorporated into media at concentrations corresponding to water activity (a(w)) values in the range 1 to 0.75. Generally, as temperature increased there was a decrease in the a(w) value at which optimum growth occurred. The a(w) limit for growth on KCl, glycerol, glucose, sorbitol, or polyethylene glycol 400 media was about 0.85, regardless of temperature. However, the a(w) limit for growth on ethanol media varied between 0.97 and 0.99 a(w) and was temperature-dependent. Water stress accounted for up to 31, 18 and 6% of growth inhibition by ethanol at 25, 40, and 42.5°C, respectively. For media containing ethanol, the decrease in growth rate per unit of a(w) reduction was greater as temperature increased. However, ethanol-induced water stress remained constant regardless of temperature, suggesting that other inhibitory effects of ethanol are closely temperature- dependent. Water stress may account for considerably more than 30% of growth inhibition by ethanol in cells that remain metabolically active at higher ethanol concentrations.

A rapid HPLC protocol for detection of polyols and trehalose

A procedure was developed to extract polyols and trehalose (protectants against stress) from fungal conidia. Conidia were sonicated (120 s) and immersed in a boiling water bath (5.5 min) to optimize extraction of polyols and trehalose, respectively. A rapid method was developed to separate and detect low-molecular-weight polyols and trehalose using high-performance liquid chromatography (HPLC). An ion exchange column designed for standard carbohydrate analysis was used in preference to one designed for sugar alcohol separation. This resulted in rapid elution (less than 5 min), without sacrificing peak resolution. The use of a pulsed electrochemical detector (gold electrode) resulted in limits of reliable quantification as low as 1.6 μg ml-1 for polyols and 2.8 μg ml-1 for trehalose. This is very sensitive and rapid method by which these protectants can be analysed. It avoids polyol derivatization that characterizes analysis by gas chromatography and the long run times (up to 45 min) that typify HPLC analysis using sugar alcohol columns.