Optimised chronic infection models demonstrate that siderophore ‘cheating’ in Pseudomonas aeruginosa is context specific

The potential for siderophore mutants of Pseudomonas aeruginosa to attenuate virulence during infection, and the possibility of exploiting this for clinical ends, have attracted much discussion. This has largely been based on the results of in vitro experiments conducted in iron-limited growth medium, in which siderophore mutants act as social ‘cheats:’ increasing in frequency at the expense of the wild type to result in low-productivity, low-virulence populations dominated by mutants. We show that insights from in vitro experiments cannot necessarily be transferred to infection contexts. First, most published experiments use an undefined siderophore mutant. Whole-genome sequencing of this strain revealed a range of mutations affecting phenotypes other than siderophore production. Second, iron-limited medium provides a very different environment from that encountered in chronic infections. We conducted cheating assays using defined siderophore deletion mutants, in conditions designed to model infected fluids and tissue in cystic fibrosis lung infection and non-healing wounds. Depending on the environment, siderophore loss led to cheating, simple fitness defects, or no fitness effect at all. Our results show that it is crucial to develop defined in vitro models in order to predict whether siderophores are social, cheatable and suitable for clinical exploitation in specific infection contexts

Comparative genomics of Pseudomonas fluorescens subclade III strains from human lungs

Abstract Background While the taxonomy and genomics of environmental strains from the P. fluorescens species-complex has been reported, little is known about P. fluorescens strains from clinical samples. In this report, we provide the first genomic analysis of P. fluorescens strains in which human vs. environmental isolates are compared. Results Seven P. fluorescens strains were isolated from respiratory samples from cystic fibrosis (CF) patients. The clinical strains could grow at a higher temperature (>34 °C) than has been reported for environmental strains. Draft genomes were generated for all of the clinical strains, and multi-locus sequence analysis placed them within subclade III of the P. fluorescens species-complex. All strains encoded type- II, −III, −IV, and -VI secretion systems, as well as the widespread colonization island (WCI). This is the first description of a WCI in P. fluorescens strains. All strains also encoded a complete I2/PfiT locus and showed evidence of horizontal gene transfer. The clinical strains were found to differ from the environmental strains in the number of genes involved in metal resistance, which may be a possible adaptation to chronic antibiotic exposure in the CF lung. Conclusions This is the largest comparative genomics analysis of P. fluorescens subclade III strains to date and includes the first clinical isolates. At a global level, the clinical P. fluorescens subclade III strains were largely indistinguishable from environmental P. fluorescens subclade III strains, supporting the idea that identifying strains as ‘environmental’ vs ‘clinical’ is not a phenotypic trait. Rather, strains within P. fluorescens subclade III will colonize and persist in any niche that provides the requirements necessary for growth.http://deepblue.lib.umich.edu/bitstream/2027.42/116129/1/12864_2015_Article_2261.pd

Unconjugated estriol in serum as measured by liquid chromatography with electrochemical detection, compared with radioimmunoassay.

Abstract Concentrations of unconjugated estriol in maternal serum or plasma increase throughout pregnancy, particularly during the third trimester. We present and discuss the results of a comparative study of unconjugated estriol as measured by a new "high-performance" liquid-chromatographic assay and by a conventional semiautomated radioimmunoassay procedure. In the new method, serum is extracted and chromatographed on a reversed-phase mu Bondapak-C8 column under radial compression. The estriol peak is detected with a glassy-carbon electrochemical detector. The chromatographic results (y) correlated well with the RIA results (x) for 105 samples from 45 pregnant women in their third trimester (y = 1.07x - 0.55 micrograms/L, r = 0.933), with no significant difference between the means of the two methods.</jats:p

Liquid-chromatographic determination of acetaminophen in serum.

Abstract We describe a sensitive and precise "high-pressure" liquid-chromatographic method for determining acetaminophen in serum. The 2-acetaminophenol and 3-acetaminophenol structural isomers of acetaminophen are used as internal standards. The method, which involves solvent extraction and adsorption chromatography on silica, provides excellent sensitivity, accuracy, and selectivity. The standard curve is linear over the range of acetaminophen concentrations of 0.5 to 300 mg/L, which makes the method useful for both pharmacokinetic studies and overdose monitoring. Analytical recovery is 97% for acetaminophen concentrations ranging from 5 to 300 mg/L. Many commonly used drugs were tested and found not to interfere. The procedure has been successfully adapted as a microscale method requiring only 50 microL of sample. The microscale method is particularly useful for pediatric and neonatal patients for whom sample size is a major concern.</jats:p

Atomic Absorption Spectrometry of Nickel, Copper, Zinc, and Lead in Sweat Collected from Healthy Subjects during Sauna Bathing

Abstract Ni, Cu, Zn, and Pb were measured by atomic absorption spectrometry in sweat samples obtained by the arm-bag technique from 48 healthy adult subjects (33 ♂, 15 ♀ ) during sauna bathing (15 min at 93 °C, dry heat). The men sweated more profusely than the women (volume, in milliliters, of sweat collected: mean, SD, and range: 23 ± 12 (3-55) and 7 ± 3 (2-13), respectively. The concentrations, in µg/liter, (mean, SD, and range) of trace metals in sweat of men and women, respectively, were: nickel, 52 ± 36 (7-180) and 131 ± 65 (39-270); copper, 550 ± 350 (30-1440) and 1480 ± 610 (590-2280); zinc, 500 ± 480 (130-1460) and 1250 ± 770 (530-2620); and lead, 51 ± 42 (8-184) and 118 ± 72 (49-283). In sweat samples from 11 women on oral contraceptives, concentrations of Ni, Cu, Zn, and Pb did not differ significantly from the values in the 15 control women. Sweating is a demonstrably significant route for excretion of trace metals, and sweating may play a role in trace-metal homeostasis. Essential trace metals could conceivably be depleted during prolonged exposure to heat; conversely, sauna bathing might provide a therapeutic method to increase elimination of toxic trace metals.</jats:p