Inhibition of bacterial growth by granulocytes measured by an automated technique

Granulocytes, collected by several methods, were assayed for antibacterial activity utilizing a technique originally developed for automated antibiotic susceptibility testing. The granulocytes were incubated with either Escherichia coli or Staphylococcus for one hour at 37 degrees C and were then separated from the suspension by gentle centrifugation at 170 g. The bacteria remaining in the supernatant broth were incubated in culture medium (eugonic broth) and their growth density was compared with diluted controls (without granulocytes) by measurement in an Autobac I (an automated nephelometer). Measurements of the density of growth at 30 minute intervals showed a marked delay in the development of density suggesting only small numbers of organisms remained after incubation with granulocytes. Once density was measurable, the rate of growth appeared similar to controls. After 2.5 hours in the Atuobac I, the density of growth was used to determine the inhibitory effect of granulocytes. The density of growth was inversely related to the concentration of granulocytes present during the preincubation phase. Filtered granulocytes showed significantly lower inhibitory effect than centrifuged granulocytes. This rapid, inexpensive method of determining microbial growth appears to be adaptable as a measure of granulocyte function.