THE GENETIC-BASIS OF COLONY OPACITY IN STREPTOCOCCUS-PNEUMONIAE - EVIDENCE FOR THE EFFECT OF BOX ELEMENTS ON THE FREQUENCY OF PHENOTYPIC VARIATION

Streptococcus pneumoniae undergoes spontaneous phase variation in colo ny morphology. Differences in colony opacity have previously been show n to correlate with differences in the ability of organisms to coloniz e the mucosal surface of the nasopharynx in an animal model. The genet ic basis of opacity variation was identified in transformation experim ents. A DNA library, from a strain that varies at high frequency, was screened to identify a single clone capable of transforming a transpar ent recipient strain which varies at low frequency to an opaque phenot ype. Analysis of this opacity locus revealed two genes, glpD and glpF, with similarity to genes required for glycerol metabolism in other ba cteria. Following the pneumococcal glpF, repetitive intergenic element s, boxes A and C, were identified. These stem-loop-forming elements we re not present in the same locus of the recipient strain. Although not required for phase variation in colony opacity, the box element was n ecessary for expression of phase variation at high frequency. Introduc tion of the box elements during transformation affected colony morphol ogy, possibly by altering expression of a putative regulatory gene dow nstream from the box element. Mutagenesis within this region confirmed the contribution of the putative regulatory gene to the expression of colony opacity. Growth characteristics of strains generated in this s tudy provide additional evidence for an association of differences in cell wall autolysis and variation in colony opacity.