The addition of Congo red, Trypan blue or haemin to the growth medium allowed the differentiation of phase-I and variant strains of Bordetella pertussis. Phase-I strains produced red (CR+), blue or dark brown colonies on a modified cyclodextrin solid medium containing Congo red, Trypan blue or haemin, respectively, whereas variant (Vir- and phase IV) strains grew as pale (CR-) colonies. Spontaneous CR- variants were isolated and characterised and had a phenotype like that of Vir- or phenotypically modulated, C-mode strains in that they did not produce the haemolysin, haemagglutinin(s), histamine-sensitising factor (pertussis toxin), heat-labile toxin and two major envelope polypeptides associated with phase-I strains. Two such variants had reduced virulence for mice. CR+ strains, when grown on a high nicotinic acid medium to induce modulation, gave CR- colonies. Thus the CR+ phenotype is a characteristic of phase-I B. pertussis and its expression appears to be controlled in a manner similar to that of other phase I-related factors. CR- variants of B. parapertussis and B. bronchiseptica were also deficient in these factors. Four isolates of B. avium were CR-.