IN-VITRO SUSCEPTIBILITIES OF BARTONELLA-HENSELAE, B-QUINTANA, B-ELIZABETHAE, RICKETTSIA-RICKETTSII, R-CONORII, R-AKARI, AND R-PROWAZEKII TOMACROLIDE ANTIBIOTICS AS DETERMINED BY IMMUNOFLUORESCENT-ANTIBODY ANALYSIS OF INFECTED VERO CELL MONOLAYERS

The in vitro susceptibilities of Bartonella (Rochalimaea) henselae, B. quintana, B, elizabethae, Rickettsia akari, R. conorii, R, prowazekii , and R, rickettsii to different concentrations of azithromycin, clari thromycin, dirithromycin, erythromycin, and roxithromycin in Vero cell cultures were evaluated, Bartonella and Rickettsia spp. were allowed to initiate infection of the antibiotic-free Vero cell monolayers, whi ch were maintained in 16-chamber microscope slides in the absence of a ntibiotics at 32 degrees C in a CO2-enriched atmosphere, The monolayer s were then incubated for 3 h to allow for initial host cell intracell ular penetration by infecting species. After inoculation, inocula were replaced and tested with media containing 12 different concentrations of each antibiotic in replicate (10 wells of each antibiotic dilution ) for each species, and the monolayers were reincubated, Tetracycline served as the control, Growth status of Bartonella spp. and Rickettsia spp, was determined by evaluation of immunofluorescent staining bacil li, Five days later, when antibiotic-free, control-infected cell monol ayers demonstrated significant fluorescence, media were removed for al l cell monolayers, the monolayers were fixed, and all specimens were s tained with standard indirect immunofluorescent antibody reagents, Flu orescent foci were enumerated by counting such foci on random fields v isualized with an epifluorescence microscope, The extent of antibiotic -induced focus inhibition was recorded for each dilution of antibiotic and compared with that of an antibiotic-negative control, Effective a ntibiotic dilution endpoints for inhibition of Bartonella and Ricketts ia proliferation, as judged by absence of increase of significant fluo rescence (as compared with no-growth controls), were enumerated by det ermining the number of cell culture chambers at various antibiotic dil utions that were negative or positive for significant Bartonella- or R ickettsia-specific fluorescence. All of the macrolide agents tested we re readily active against all three Bartonella organisms, and azithrom ycin, clarithromycin, and roxithromycin may have potential in the trea tment of Rickettsia infections, Animal model-based clinical trials are warranted to define the specific treatment role of the newer macrolid e antibiotics.