Browsing by Author "Pettigrew, Charles"

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Imaging and plate counting to quantify the effect of an antimicrobial: A case study of a photo-activated chlorine dioxide treatment
(Oxford University Press, 2022-12) Parker, Albert E.; Miller, Lindsey; Adams, Jacob; Pettigrew, Charles; Buckingham-Meyer, Kelli; Summers, Summers; Christen, Andres; Goeres, Darla
Aim. To assess removal versus kill efficacies of antimicrobial treatments against thick biofilms with statistical confidence. Methods and results. A photo‐activated chlorine dioxide treatment (Photo ClO2) was tested in two independent experiments against thick (>100 μm) Pseudomonas aeruginosa biofilms. Kill efficacy was assessed by viable plate counts. Removal efficacy was assessed by 3D confocal scanning laser microscope imaging (CSLM). Biovolumes were calculated using an image analysis approach that models the penetration limitation of the laser into thick biofilms using Beer's Law. Error bars are provided that account for the spatial correlation of the biofilm's surface. The responsiveness of the biovolumes and plate counts to the increasing contact time of Photo ClO2 were quite different, with a massive 7 log reduction in viable cells (95% confidence interval [CI]: 6.2, 7.9) but a more moderate 73% reduction in biovolume (95% CI: [60%, 100%]). Results are leveraged to quantitatively assess candidate CSLM experimental designs of thick biofilms. Conclusions. Photo ClO2 kills biofilm bacteria but only partially removes the biofilm from the surface. To maximize statistical confidence in assessing removal, imaging experiments should use fewer pixels in each z‐slice, and more importantly, at least two independent experiments even if there is only a single field of view in each experiment. Significance and impact of study. There is limited penetration depth when collecting 3D confocal images of thick biofilms. Removal can be assessed by optimally fitting Beer's Law to all of the intensities in a 3D image and by accounting for the spatial correlation of the biofilm's surface. For thick biofilms, other image analysis approaches are biased or do not provide error bars. We generate unbiased estimates of removal and assess candidate CSLM experimental designs of thick biofilms with different pixilations, numbers of fields of view and number of experiments using the included design tool.
Imaging and plate counting to quantify the effect of an antimicrobial: A case study of a photo‐activated chlorine dioxide treatment
(Wiley, 2022-09) Parker, Albert E.; Miller, Lindsey; Adams, Jacob; Pettigrew, Charles; Buckingham‐Meyer, Kelli; Summers, Jennifer; Christen, Andres; Goeres, Darla
Aim. To assess removal versus kill efficacies of antimicrobial treatments against thick biofilms with statistical confidence. Methods and results. A photo-activated chlorine dioxide treatment (Photo ClO2) was tested in two independent experiments against thick (>100 μm) Pseudomonas aeruginosa biofilms. Kill efficacy was assessed by viable plate counts. Removal efficacy was assessed by 3D confocal scanning laser microscope imaging (CSLM). Biovolumes were calculated using an image analysis approach that models the penetration limitation of the laser into thick biofilms using Beer's Law. Error bars are provided that account for the spatial correlation of the biofilm's surface. The responsiveness of the biovolumes and plate counts to the increasing contact time of Photo ClO2 were quite different, with a massive 7 log reduction in viable cells (95% confidence interval [CI]: 6.2, 7.9) but a more moderate 73% reduction in biovolume (95% CI: [60%, 100%]). Results are leveraged to quantitatively assess candidate CSLM experimental designs of thick biofilms. Conclusions. Photo ClO2 kills biofilm bacteria but only partially removes the biofilm from the surface. To maximize statistical confidence in assessing removal, imaging experiments should use fewer pixels in each z-slice, and more importantly, at least two independent experiments even if there is only a single field of view in each experiment. Significance and impact of study There is limited penetration depth when collecting 3D confocal images of thick biofilms. Removal can be assessed by optimally fitting Beer's Law to all of the intensities in a 3D image and by accounting for the spatial correlation of the biofilm's surface. For thick biofilms, other image analysis approaches are biased or do not provide error bars. We generate unbiased estimates of removal and assess candidate CSLM experimental designs of thick biofilms with different pixilations, numbers of fields of view and number of experiments using the included design tool.
Integrated molecular, physiological and in silico characterization of two Halomonas isolates from industrial brine
(2016-05) Carlson, Ross P.; Oshota, Olusegun J.; Shipman, Matt R.; Caserta, J. A.; Hu, P.; Saunders, C. W.; Xu, Jun; Jay, Zackary J.; Reeder, N.; Richards, Abigail M.; Pettigrew, Charles; Peyton, Brent M.
Two haloalkaliphilic bacteria isolated from industrial brine solutions were characterized via molecular, physiological, and in silico metabolic pathway analyses. Genomes from the organisms, designated Halomonas BC1 and BC2, were sequenced; 16S ribosomal subunit-based phylogenetic analysis revealed a high level of similarity to each other and to Halomonas meridiana. Both strains were moderate halophiles with near optimal specific growth rates (=60 % Âµmax) observed over <0.1â€“5 % (w/v) NaCl and pH ranging from 7.4 to 10.2. Isolate BC1 was further characterized by measuring uptake or synthesis of compatible solutes under different growth conditions; in complex medium, uptake and accumulation of external glycine betaine was observed while ectoine was synthesized de novo in salts medium. Transcriptome analysis of isolate BC1 grown on glucose or citrate medium measured differences in glycolysis- and gluconeogenesis-based metabolisms, respectively. The annotated BC1 genome was used to build an in silico, genome-scale stoichiometric metabolic model to study catabolic energy strategies and compatible solute synthesis under gradients of oxygen and nutrient availability. The theoretical analysis identified energy metabolism challenges associated with acclimation to high salinity and high pH. The study documents central metabolism data for the industrially and scientifically important haloalkaliphile genus Halomonas.