Phenotypic variation in Haemophilus influenzae: the interrelationship of colony opacity, capsule and lipopolysaccharide.

H. influenzae type b strains show phase variation between opaque (O) and translucent (T) colony phenotypes. These phenotypic differences have been related to differences in virulence for infant rats. This study shows that the switch between O and T colony phenotypes is associated with variation in the amount of cell-associated capsule in the serotype b strains Rd:b+:01, RM7004 and Eagan. O colonies comprised organisms which were more serum resistant and had more cell-associated polyribosyl ribitol phosphate (PRP) than organisms from T colonies. Strain Rd, the non-encapsulated parent of the encapsulated transformant Rd:b+:01, was constitutively translucent, consistent with its lack of capsule expression. Since previous studies had correlated O-T switching with differences in the relative molecular weight of lipopolysaccharide (LPS), LPS phenotypes of Rd and Rd:b+:01 were compared and correlated with opacity phenotype at the individual colony level. Both strains showed phase variation between higher and lower molecular weight LPS oligosaccharide structures but the prevalence of higher molecular weight LPS was greater for the capsule-deficient Rd than encapsulated Rd:b+01. Capsule-deficient mutants of strains Rd:b+:01, RM7004 and Eagan produced constitutively translucent colonies and each had a greater prevalence of higher molecular weight LPS than their encapsulated parents. These findings indicated an incomplete association between capsular O-T phase variation and LPS expression.