RICKETTSIA-TSUTSUGAMUSHI INFECTION IN CELL-CULTURE - ANTIBIOTIC SUSCEPTIBILITY DETERMINED BY FLOW-CYTOMETRY

Recent unpublished reports from northern Thailand of severe and someti mes fatal cases of scrub typhus, despite appropriate antibiotic therap y, suggest that resistance may occur. Current antibiotic susceptibilit y methods that use direct microscopic counts of Giemsa-stained cells o r mouse protection assays are slow, labor-intensive, and expensive. We explored the use of flow cytometry to measure rickettsial infection i n vitro in L-929 cells treated with and without doxycycline, ciproflox acin, erythromycin, and chloramphenicol. It was possible to detect the rickettsiae down to a level of 83% of the cells infected, mean of 37 rickettsiae per cell, and 40% of cells with too many rickettsiae to co unt. This level of sensitivity was sufficient to determine the inhibit ory effect of all four drugs at standard screening concentrations. At lower concentrations of doxycycline, flow cytometry detected inhibitio n of rickettsial growth at a concentration of 6.25 X 10(-2) mu g/ml bu t not at 6.25 X 10(-3) mu g/ml, suggesting that the minimum inhibitory concentration is somewhere between these two values. The data from th is study show that Row cytometry permits the rapid screening of numero us rickettsial isolates for their susceptibility to a variety of antib iotics, but that visual counts of infected cells provide a more precis e indication of rickettsial growth.