Morphology and viability of large bodies of streptococcal L-forms

A procedure is described for the massive formation and isolation of large bodies of group A and D streptococcal L-forms. Up to 90% individual bodies of 20 to 100 mum in diameter can be produced in pour plates of nutrient gelatin and subsequently harvested by simple micromanipulation. The growth of these giant bodies was followed by light microscopy and their ultrastructure and internal architecture was examined by transmission and scanning electron microscopy. Large bodies had a honeycomb structure of vacuoles. Numerous small units could be observed inside, protruding into, and between vacuoles. Intact large bodies (30 to 60 mum), when placed in small amounts of broth and incubated at 35 C, initiated turbid cultures, but when set on agar, they ruptured, releasing internal granules and producing typical L-form colonies. The number of internal colony-forming units (CFU) was correlated with the size of large bodies. Up to 200 CFU were detected in bodies of 40 to 60 mum in diameter, whereas corpuscles of 20 to 30 mum averaged only three CFU. The inefficiency of replication was apparent with the determination by light and electron microscopy that at least 100 times as many granules and elementary corpuscles as CFU were produced inside large bodies.