DNA replication and cell enlargement of Enterococcus faecalis protoplasts

Protoplasts of Enterococcus faecalis did not divide but enlarged in Difco Marine Broth containing penicillin. Our previous studies have demonstrated that transcription and translation were essential for bacterial cell enlargement. However, it was uncertain whether replication was also essential. In this study, we measured the amount of DNA in E. faecalis cells during the course of enlargement using quantitative polymerase chain reaction. The growth of normally divided cells (native forms) of E. faecalis exhibited a log phase before 6 h of incubation was reached. Although a difference in quantitation cycle (Cq) values between the replication initiation and termination regions was observed in the log phase, it was not present in the stationary growth phase. On the other hand, the amount of DNA in E. faecalis protoplasts increased during the cell enlargement incubation. The difference of Cq values between the protoplasts at 0 and 96 h of incubation was 8-9, indicating that the DNA amount at 96 h was 200-500 times higher than that at 0 h. The Cq values differed between the replication initiation and termination regions, indicating that the replication level was high. When novobiocin, a DNA replication inhibitor, was added to the medium at 24 h of incubation, DNA replication and cell enlargement were almost stopped. Thus, replication plays an important role in the enlargement of E. faecalis protoplasts.