Browsing by Author "Goeres, Darla"

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Evaluation of 3M Petrifilm as an equivalent alternative to drop-plating on agar plates in a biofilm system
(2013-03) Fritz, Blaine; Goeres, Darla
This project evaluated 3M Petrifilm as an alternative, more efficient method for bacterial enumeration. Using Petrifilm allows the researcher to avoid preparing agar plates for bacterial enumeration. Currently, the majority of scientific literature concerning enumeration of bacteria on Petrifilm is from the food industry. There are no published studies examining the use of Petrifilm for enumeration of biofilm bacteria. A Pseudomonas aeruginosa biofilm was grown in a CDC reactor according to ASTM Method E2562. The mature biofilm was exposed to chlorine (buffered water for controls) and neutralized. The biofilm was removed from the surface, disaggregated, and serially diluted. Samples from the dilution tubes were plated in duplicate on Petrifilm Aerobic Count plates and drop plated on R2A plates. The Petrifilm and R2A plates were incubated at 36oC and colonies enumerated after 24 and 48 hours. The experiment was replicated three times by two technicians. The time required for both plating methods was recorded to help assess the efficiency of both methods. The results from this study may demonstrate that Petrifilm could replace drop plating as a more efficient and cost effective method for bacterial enumeration.
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.