Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical Confidence
The goal of this study was to quantify the variability of confocal laser scanning microscopy (CLSM) time-lapse images of early colonizing biofilms to aid in the design of future imaging experiments. To accomplish this a large imaging dataset consisting of 16 independent CLSM microscopy experiments w...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Frontiers Media S.A.
2022-01-01
|
| Series: | Frontiers in Microbiology |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2021.785182/full |
| _version_ | 1798021303389650944 |
|---|---|
| author | Brian A. Pettygrove Brian A. Pettygrove Heidi J. Smith Heidi J. Smith Kyler B. Pallister Jovanka M. Voyich Philip S. Stewart Philip S. Stewart Albert E. Parker Albert E. Parker |
| author_facet | Brian A. Pettygrove Brian A. Pettygrove Heidi J. Smith Heidi J. Smith Kyler B. Pallister Jovanka M. Voyich Philip S. Stewart Philip S. Stewart Albert E. Parker Albert E. Parker |
| author_sort | Brian A. Pettygrove |
| collection | DOAJ |
| description | The goal of this study was to quantify the variability of confocal laser scanning microscopy (CLSM) time-lapse images of early colonizing biofilms to aid in the design of future imaging experiments. To accomplish this a large imaging dataset consisting of 16 independent CLSM microscopy experiments was leveraged. These experiments were designed to study interactions between human neutrophils and single cells or aggregates of Staphylococcus aureus (S. aureus) during the initial stages of biofilm formation. Results suggest that in untreated control experiments, variability differed substantially between growth phases (i.e., lag or exponential). When studying the effect of an antimicrobial treatment (in this case, neutrophil challenge), regardless of the inoculation level or of growth phase, variability changed as a frown-shaped function of treatment efficacy (i.e., the reduction in biofilm surface coverage). These findings were used to predict the best experimental designs for future imaging studies of early biofilms by considering differing (i) numbers of independent experiments; (ii) numbers of fields of view (FOV) per experiment; and (iii) frame capture rates per hour. A spreadsheet capable of assessing any user-specified design is included that requires the expected mean log reduction and variance components from user-generated experimental results. The methodology outlined in this study can assist researchers in designing their CLSM studies of antimicrobial treatments with a high level of statistical confidence. |
| first_indexed | 2024-04-11T17:11:30Z |
| format | Article |
| id | doaj.art-7d5a8de0a6b84dc2919fb04ebe45d091 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-04-11T17:11:30Z |
| publishDate | 2022-01-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-7d5a8de0a6b84dc2919fb04ebe45d0912022-12-22T04:12:54ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2022-01-011210.3389/fmicb.2021.785182785182Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical ConfidenceBrian A. Pettygrove0Brian A. Pettygrove1Heidi J. Smith2Heidi J. Smith3Kyler B. Pallister4Jovanka M. Voyich5Philip S. Stewart6Philip S. Stewart7Albert E. Parker8Albert E. Parker9Center for Biofilm Engineering, Montana State University, Bozeman, MT, United StatesDepartment of Microbiology and Cell Biology, Montana State University, Bozeman, MT, United StatesCenter for Biofilm Engineering, Montana State University, Bozeman, MT, United StatesDepartment of Microbiology and Cell Biology, Montana State University, Bozeman, MT, United StatesDepartment of Microbiology and Cell Biology, Montana State University, Bozeman, MT, United StatesDepartment of Microbiology and Cell Biology, Montana State University, Bozeman, MT, United StatesCenter for Biofilm Engineering, Montana State University, Bozeman, MT, United StatesDepartment of Chemical and Biological Engineering, Montana State University, Bozeman, MT, United StatesCenter for Biofilm Engineering, Montana State University, Bozeman, MT, United StatesDepartment of Mathematical Sciences, Montana State University, Bozeman, MT, United StatesThe goal of this study was to quantify the variability of confocal laser scanning microscopy (CLSM) time-lapse images of early colonizing biofilms to aid in the design of future imaging experiments. To accomplish this a large imaging dataset consisting of 16 independent CLSM microscopy experiments was leveraged. These experiments were designed to study interactions between human neutrophils and single cells or aggregates of Staphylococcus aureus (S. aureus) during the initial stages of biofilm formation. Results suggest that in untreated control experiments, variability differed substantially between growth phases (i.e., lag or exponential). When studying the effect of an antimicrobial treatment (in this case, neutrophil challenge), regardless of the inoculation level or of growth phase, variability changed as a frown-shaped function of treatment efficacy (i.e., the reduction in biofilm surface coverage). These findings were used to predict the best experimental designs for future imaging studies of early biofilms by considering differing (i) numbers of independent experiments; (ii) numbers of fields of view (FOV) per experiment; and (iii) frame capture rates per hour. A spreadsheet capable of assessing any user-specified design is included that requires the expected mean log reduction and variance components from user-generated experimental results. The methodology outlined in this study can assist researchers in designing their CLSM studies of antimicrobial treatments with a high level of statistical confidence.https://www.frontiersin.org/articles/10.3389/fmicb.2021.785182/fullmicroscopybiofilmimage analysisstatistical confidencerepeatabilityexperimental design |
| spellingShingle | Brian A. Pettygrove Brian A. Pettygrove Heidi J. Smith Heidi J. Smith Kyler B. Pallister Jovanka M. Voyich Philip S. Stewart Philip S. Stewart Albert E. Parker Albert E. Parker Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical Confidence Frontiers in Microbiology microscopy biofilm image analysis statistical confidence repeatability experimental design |
| title | Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical Confidence |
| title_full | Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical Confidence |
| title_fullStr | Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical Confidence |
| title_full_unstemmed | Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical Confidence |
| title_short | Experimental Designs to Study the Aggregation and Colonization of Biofilms by Video Microscopy With Statistical Confidence |
| title_sort | experimental designs to study the aggregation and colonization of biofilms by video microscopy with statistical confidence |
| topic | microscopy biofilm image analysis statistical confidence repeatability experimental design |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2021.785182/full |
| work_keys_str_mv | AT brianapettygrove experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT brianapettygrove experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT heidijsmith experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT heidijsmith experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT kylerbpallister experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT jovankamvoyich experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT philipsstewart experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT philipsstewart experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT alberteparker experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence AT alberteparker experimentaldesignstostudytheaggregationandcolonizationofbiofilmsbyvideomicroscopywithstatisticalconfidence |