Real time computer tracking of free-swimming and tethered rotating cells

A computerized image processing system has been developed that tracks individual free-swimming cells and rotating bacterial cell bodies tethered by their flagella in real time. Free-swimming bacteria of Rhodobacter sphaeroides, Rhodospirullum rubrum, and Salmonella typhimurium have been tracked swimming at speeds from 0 to over 120 microns s-1. A high level of discrimination is exerted against noncellular objects, allowing analysis of stopped as well as moving cells. This enabled detection of both speed and qualitative change in the swimming patterns of R. sphaeroides WS8 upon tactic stimulation. Comparison with darkfield microscopy indicated that the two techniques were in substantial agreement. The unidirectional rotation of cells of R. sphaeroides WS8 could be detected when the cells were either parallel to the microscope slide or end on. Frequencies of rotation of up to 10 Hz were monitored before image blurring became a problem. True rods would be easier to analyze at higher speeds of rotation. Although developed for photosynthetic bacteria, a wide range of bacteria, eucaryotic organisms, and subcellular organelles could be tracked with this system. Minor modifications to the software allow customization to different types of motility analysis.