USE OF COMPUTER-ASSISTED MOTION ANALYSIS FOR QUANTITATIVE MEASUREMENTS OF SWIMMING BEHAVIOR IN PERITRICHOUSLY FLAGELLATED BACTERIA

An assay was developed which identifies individual bacterial tumbles a nd so allows rapid, quantitative measurements of tumble frequency in f ree-swimming bacteria. Tumble frequency is modulated by cells to enabl e chemotaxis. Mutations in the chemotaxis signal transduction pathway typically have phenotypes of altered tumble frequency. The purpose of this assay is to quantitatively measure steady-state tumble frequency to enable comparisons of mutant strain phenotypes. It was developed us ing Escherichia coli but should be applicable to other species with a peritrichous flagellation pattern, such as Salmonella typhimurium. Tum bles are defined by a combination of the parameters rate of change of direction and swimming speed, with a rapid change of direction definin g the beginning of a tumble and increased swimming speed defining the end. These parameters have previously been shown to be correlated with tumbles in general but not used to identify discrete tumble events. T he computer assay was validated by comparing its results with manual o bservations by eye. The assay was intended to be most sensitive to swi mming patterns similar to wild type so as to resolve subtle changes wh ich would result from partial-function mutations. It quantitatively de tects extreme behavioral phenotypes as well and can be modified to inc rease resolution at either extreme if necessary. (C) 1996 Academic Pre ss, Inc.