IMPROVED PLAQUE ASSAYS FOR RICKETTSIA-PROWAZEKII IN VERO76 CELLS

Typhus group rickettsiae, including Rickettsia prowazekii and R. typhi , produce visible plaques on primary chick embryo fibroblasts and low- passage mouse embryo fibroblasts but do not form reproducible plaques on continuous cell culture lines, We tested medium overlay modificatio ns for plaque formation of typhus group rickettsiae on the continuous fibroblast cell line Vero76, A procedure involving primary overlay wit h medium at pH 6.8, which was followed 2 to 3 days later with secondar y overlay at neutral pH containing 1 mu g of emetine per ml and 20 mu g of NaF per ml, resulted in visible plaques at 7 to 10 days postinfec tion, A single-step procedure involving overlay with medium containing 50 ng of dextran sulfate per ml also resulted in plaque formation wit hin 8 days postinfection, These assays represent reproducible and inex pensive methods for evaluating the infectious titers of typhus group r ickettsiae, cloning single plaque isolates, and testing the susceptibi lities of rickettsiae to antibiotics.