Evaluation of the Accuracy and Precision of Enumerating Aerobic Heterotrophs in Water Samples by the Spread Plate Method

Abstract

Factors associated with accuracy and precision in the enumeration of aquatic aerobic heterotrophs by the spread plate method were evaluated by using a nested analysis of variance experimental design. Variances associated with individual components of the spread plate procedure were isolated, and optimal replications of each step were allocated. A practical scheme for optimal allocation of resources is proposed, consisting of four subsamples and two plates per subsample and yielding a total variance decrease of 70% from a single-subsample, 10-plate series. Data transformation was, in general, unnecessary for intraexperiment or intrasample statistical analysis, whereas interexperiment or intersample comparisons may require transformation of data. Rapid changes in the numbers of organisms in stored water samples were observed that were not reproducible and did not follow detectable trends, with increases or decreases in counts occurring in samples regardless of whether they were stored at room temperature or refrigerated, or stored in plastic or glass containers. Rapid sample handling is strongly recommended to minimize variations in the microbial populations of samples for aquatic environments.