The production and regeneration of bacterial protoplasts promoted the loss of three different plasmid-specified traits in Streptococcus lactis subsp. diacetylactis strains. The loss of five different plasmids, including small multicopy molecules, was readily detected in Streptococcus lactis 712 by screening lysates of random protoplast regenerants on agarose gels. In this strain sequential rounds of protoplast regeneration were used to produce a plasmid-free strain and derivatives carrying only single molecules from the plasmid complement. During these experiments a 33-megadalton plasmid, pLP712, was found to encode genes for lactose and protein utilization. Only this plasmid was required for normal growth and acid production in milk; the remaining four plasmids appeared to be cryptic. Lactose-defective derivatives of a strain carrying only pLP712 were readily isolated. Although these derivatives included instances of plasmid loss, deletions of pLP712 were frequently found. Many different deleted derivatives of pLP712, including some in which the lactose or protein utilization determinant or both were lost, were isolated. The molecular instability of pLP712 largely accounted

Plasmid DNA from lactic streptococci was subjected to electrophoresis on agarose gels. The plasmid profiles so obtained were strain specific and sufficiently stable to suggest their use in strain differentiation. A group of Streptococcus lactis strains, 712. 763 (ML3), 505 (C2) and 2031 (C2), found to have similar plasmid profiles, were shown to be closely related. Gene transfer by transduction and conjugation occurred between members of this group at frequencies comparable to those in homologous systems and temperate phages cross plated readily between their prophage cured derivatives. Minor variations were, however, found between these four strains; slight differences in plasmid profiles, lysogenic status, prophage curability and temperate phage morphology were detected and it is suggested that these have evolved as a result of maintenance in different environments.

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The broad-host-range plasmid pAMβ1, which codes for erythromycin and lincomycin resistance, was transferred by conjugation into Lactobacillus acidophilus, Lactobacillus reuteri , and Lactobacillus salivarius. A novel 17-megadalton plasmid molecule was detected in the transconjugants, confirming the introduction of pAMβ1 into each species.

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