GENERATION AND PROPERTIES OF A STREPTOCOCCUS-PNEUMONIAE MUTANT WHICH DOES NOT REQUIRE CHOLINE OR ANALOGS FOR GROWTH

A mutant (JY2190) of Streptococcus pneumoniae Rx1 which had acquired t he ability to gravy in the absence of choline and analogs was isolated . Lipoteichoic acid (LTA) and wall teichoic acid (TA) isolated from th e mutant were free of phosphocholine and other phosphorylated amino al cohols. Both polymers showed an unaltered chain structure and, in the case of LTA, an unchanged glycolipid anchor. The cell wall composition was also not altered except that, due to the lack of phosphocholine, the phosphate content of cell walls was half that of the parent strain . Isolated cell walls of the mutant were resistant to hydrolysis by pn eumococcal autolysin (N-acetylmuramyl-L-alanine amidase) but were clea ved by the muramidases CPL and cellosyl. The lack of active autolysin in the mutant cells became apparent by impaired cell separation at the end of cell division and by resistance against stationary-phase and p enicillin-induced lysis. As a result of the absence of choline in the LTA, pneumococcal surface protein A (PspA) was no longer retained on t he cytoplasmic membrane. During growth in the presence of choline, whi ch was incorporated as phosphocholine into LTA and TA, the mutant cell s separated normally, did not release PspA, and became penicillin sens itive. However, even under these conditions, they did not lyse in the stationary phase, and they showed poor reactivity with antibody to pho sphocholine and an increased release of C-polysaccharide from the cell . In contrast to ethanolamine-grown parent cells (A, Tomasz, Proc. Nat l, Acad, Sci. USA 59:86-93, 1968), the choline-free mutant cells retai ned the capability to undergo genetic transformation but, compared to Rx1, with lower frequency and at an earlier stage of growth. The prope rties of the mutant could be transferred to the parent strain by DNA o f the mutant.