The effect of (alpha 2-->8)-linked polysialic acid on the adherence of Neisseria meningitidis to human mucosal cells was examined using a serogroup B-encapsulated strain and a capsule-defective (Cap-) mutant of this strain. The Cap- mutant contains a single truncated insert of Tn916 in a 3.8-kb HaeIII chromosomal fragment. The Tn916 insert was shown to be responsible for the phenotype by linkage studies and by demonstration that loss of the insert restored encapsulation. The Cap- mutant consistently adhered to human buccal epithelial cell in greater numbers than the encapsulated parent, but the increase in adherence was less than twofold. Adherence of the Cap- mutant during infection of human nasopharyngeal organ cultures was 1.3- to 6.5-fold greater than that of the encapsulated parent. However, specificity of adherence of meningococci for nonciliated nasopharyngeal epithelial cells and the ability to be internalized by these cells was not due to the (alpha 2-->8)-linked polysialic acid capsule.

The difficulty in obtaining mutants in pathogenic Neisseria has limited the ability to genetically define determinants responsible for virulence as well as the ability to generate a genetic map. We show that the 16.5kb conjugative transposon Tn916 can be introduced into Neisseria meningitidis on the suicide vectors pAM120 and pAM170. After introduction, Tn916 transposed to different sites in the chromosome of recipient meningococci, apparently at random, and was stably incorporated. Following its integration into the meningococcal chromosome, Tn916 did not appear to readily express its conjugative and transpositional functions. However, chromosomal DNA from Tn916-carrying meningococci could be used to transform other meningococcal strains to tetracycline resistance. These studies indicate that Tn916 may be an important tool for genetic analysis of N. meningitidis.

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In order to investigate possible functional consequences of phase and antigenic variation of meningococci, the attachment of 15 strains of Neisseria meningitidis to human erythrocytes was studied by a nitrocellulose hemadsorption assay. This assay allows the study of individual meningococcal colonies with respect to erythrocyte attachment. Of the 15 strains studied, 7 demonstrated binding of human erythrocytes (HA').Among these seven strains, the percentage of colonies that were HA' ranged from 0.2 to 97%. Meningococcal colonies that did not produce pilin (the major structural subunit of pili) did not demonstrate erythrocyte binding (HA-). The HA+ colony phenotype was correlated with assembly of pilin into pili and expression of pili on the meningococcal surface. However, only some piliated colonies bound human erythrocytes. This could not be explained by differences between piliated HA' and HA-colonies in the amount of pilin produced or by differences in number of pili expressed per diplococcus. Pili of five of the meningococcal strains with HA' colonies were antigenically related to gonococcal pili (class I meningococcal pili), but HA' colonies were also seen in two meningococcal strains expressing class II meningococcal pili. Changes from HA' to HA-and from HAto HA+, in the presence of continuing pilin production and pilus assembly, occurred at frequencies of up to l0-2/CFU per generation. Such frequencies resemble those of phase and antigenic variation described previously for Neisseria species pilin. These studies indicate that phase variation influences the ability of meningococci to attach to human cells and suggest that meningococci may express functionally different pili.