How to optimize the drop plate method for enumerating bacteria

Herigstad, Becky; Hamilton, Martin A.; Heersink, Joanna

doi:10.1016/s0167-7012(00)00241-4

Cited by 758 publications

(567 citation statements)

References 12 publications

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“…for 60 s. The disaggregated biofilm was then processed to quantify the number of viable cells. This entailed drop-plating serial dilutions of the suspension onto R2A agar (Difco), incubating the plates at 37 uC for 24 h, and counting the colonies (Herigstad et al, 2001). …”

Section: Methodsmentioning

confidence: 99%

Statistical assessment of a laboratory method for growing biofilms

et al. 2005

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Microbial biofilms have been grown in laboratories using a variety of different approaches. A laboratory biofilm reactor system, called the CDC biofilm reactor (CBR) system, has been devised for growing biofilms under moderate to high fluid shear stress. The reactor incorporates 24 removable biofilm growth surfaces (coupons) for sampling and analysing the biofilm. Following preliminary experiments to verify the utility of the CBR system for growing biofilms of several clinically relevant organisms, a standard operating procedure for growing a Pseudomonas aeruginosa biofilm was created. This paper presents the results of a rigorous, intra-laboratory, statistical evaluation of the repeatability and ruggedness of that procedure as well as the results of the experiments with clinically relevant organisms. For the statistical evaluations, the outcome of interest was the density (c.f.u. cm "2 ) of viable P. aeruginosa. Replicate experiments were conducted to assess the repeatability of the log density outcome. The mean P. aeruginosa log 10 density was 7?1, independent of the coupon position within the reactor. The repeatability standard deviation of the log density based on one coupon per experiment was 0?59. Analysis of variance showed that the variability of the log density was 53 % attributable to within-experiment sources and 47 % attributable to between-experiments sources. The ruggedness evaluation applied response-surface design and regression analysis techniques, similar to those often used for sensitivity analyses in other fields of science and engineering. This approach provided a quantitative description of ruggedness; specifically, the amount the log density was altered by small adjustments to four key operational factors -time allowed for initial surface colonization, temperature, nutrient concentration, and fluid shear stress on the biofilm. The small size of the regression coefficient associated with each operational factor showed that the method was rugged; that is, relatively insensitive to minor perturbations of the four factors. These results demonstrate that the CBR system is a reliable experimental tool for growing a standard biofilm in the laboratory and that it can be adapted to study several different micro-organisms.

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Section: Methodsmentioning

confidence: 99%

Statistical assessment of a laboratory method for growing biofilms

et al. 2005

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“…22 The plates were incubated for 48 h at 37°C, 10% CO 2 (IG 150, Jouan incubator). CFU were counted and the results were expressed as CFU/mg of biofilm dry weight.…”

Section: Bacterial Viabilitymentioning

confidence: 99%

S. mutans biofilm model to evaluate antimicrobial substances and enamel demineralization

2010

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The aim of this study was to validate a model of S. mutans biofilm formation, which simulated 'feast-famine' episodes of exposure to sucrose that occur in the oral cavity, showed dose-response susceptibility to antimicrobials and allowed the evaluation of substances with anticaries potential. S. mutans UA159 biofilms were grown for 5 days on bovine enamel slabs at 37°C, 10% CO 2 . To validate the model, the biofilms were treated 2x/day with chlorhexidine digluconate (CHX) at 0.012, 0.024 and 0.12% (concentration with recognized anti-plaque effect) and 0.05% NaF (concentration with recognized anti-caries effect). CHX showed dose-response effect decreasing biomass, bacterial viability and enamel demineralization (p < 0.05). Whereas, 0.05% NaF did not show antimicrobial effect but had similar effect to that of 0.12% CHX decreasing enamel demineralization (p < 0.05). The model developed has potential to evaluate the effect of substances on biofilm growth and on enamel demineralization.

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“…The method used here for the determination of microbial viability by the plate count, is routine in microbiology laboratories worldwide and its accuracy with most organisms is well accepted (28). However, it is possible that the method may have influenced the study results.…”

Section: Discussionmentioning

confidence: 99%

Quantification of Streptococcus mutans in Different Types of Ligature Wires and Elastomeric Chains

et al. 2017

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This study aimed to test the hypothesis that Streptococcus mutans contamination levels differ according to the type of the orthodontic ligature. Thirteen patients were selected. Each quadrant was randomly subjected to one of the following ligature-use protocols: I) elastomeric chain, II) steel ligature crossed over the archwire, III) steel ligature crossed under the archwire, and IV) steel ligature in a figure-eight pattern under the archwire. After seven days, the devices were removed and the Streptococcus mutans colony-forming unit count per mg of biofilm weight was determined. Twelve specimens (n=3) were also processed for scanning electron microscopy analysis. ANOVA and Tukey-Kramer test were used for comparisons to assess S. mutans differences between groups at a 5% significance level. There was no statistical difference in detectable levels of S. mutans among the groups (p=0.294). Scanning electron microscopy results showed abundant biofilms and microbial contamination in all groups. In conclusion, S. mutans contamination levels are similar in the different orthodontic ligatures.

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How to optimize the drop plate method for enumerating bacteria

Cited by 758 publications

References 12 publications

Statistical assessment of a laboratory method for growing biofilms

Statistical assessment of a laboratory method for growing biofilms

S. mutans biofilm model to evaluate antimicrobial substances and enamel demineralization

Quantification of Streptococcus mutans in Different Types of Ligature Wires and Elastomeric Chains

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