Cells of Escherichia coli B infected with the immunity-negative (imm2) mutant of bacteriophage T4 are able to develop a substantial level of immunity to superinfecting phage ghosts if the ghost challenge is made late in infection. This background immunity is not seen in infections with phage carrying the spackle (s) mutation in addition to the imm2 lesion. The level of immunity in s(-) infections is intermediate between that of imm(-) and wild-type infections under standard assay conditions. With respect to genetic exclusion of superinfecting phage, cells infected with imm(-) phage are completely deficient, whereas infections with the s(-) phage are only partially deficient compared to wild-type infections. Whereas s(-)-infected cells are unable to resist lysis from without by a high multiplicity of infection (MOI) of superinfecting phage, cells infected with imm(-) phage show less than wild-type levels of resistance and the majority of cells remaining intact are unable to incorporate leucine or form infective centers. Under conditions of superinfection by low MOI of homologous phage, imm(-)-infected cells are lysis inhibited, whereas s(-)-infected cells do not show this property. Superinfecting phage inject their DNA into imm(-)-infected cells with the same efficiency as seen in wild-type infections, but this efficiency is reduced when the cells are first infected with s(-) phage. The s function of T4 appears not only to affect the host cell wall as previously postulated by Emrich, but may also affect the junctures of cell wall and membrane with consequences similar to those of the imm function.