When grown in soft agar medium with its host, a bacteriophage will form a plaque, an area of clearance in the bacterial lawn. The plaque assay is a widely used technique in bacteriophage research, and is frequently used to measure phage titers and fitness.

After a serial passaging evolution experiment, replicate lines of bacteriophage ΦX174 did not produce any plaques when plated. However, using qPCR data we confirmed that not only was phage still present in our samples, but its growth rate in liquid media increased greatly over the course of the experiment. We found that this inability to form plaques was hereditary and that growth under elevated mutation rate allowed us to isolate some revertants. We have also used a reverse genetic approach to investigate the basis for this unusual phenotype, and we discuss possible explanations for its evolution.