Heat-resistant derivatives of a 168 strain carrying an mutation, which causes heat-sensitive induction of the PBSX prophage, have been isolated and screened for the acquisition of auxotrophy. Two classes of auxotrophs were isolated, namely Pro and ProMet; they lacked the ability to produce PBSX, as shown by their resistance to mitomycin C-induced lysis. The proline and methionine requirements and the resistance to mitomycin C were shown to segregate together in phage PBS1-mediated transduction crosses and to be linked to , which is known to be co-transducible with the PBSX prophage. It was therefore proposed that these strains had deletions which removed all or part of the PBSX prophage together with adjacent bacterial DNA encoding the () and genes. The mutation was shown to be in PBS1 transduction crosses; this gene is known to be co-transducible with the PBSX prophage. The proline requirement was probably due to the deletion of a gene which was demonstrated to lie between the PBSX prophage and and which was 90% co-transducible with .

These deletions have been transduced into a strain which was cured of phage SPβ, another bacteriophage carried by 168. No phage particles could be seen in mitomycin C-induced cultures of such strains.

The PBSX-deletion strains grew with the same generation time as the PBSX parent in L-broth (27 min at 35 °C) but they were slower in minimal medium (e.g. 72 min as against 51 min in the PBSX strain). Besides being resistant to mitomycin C-induced lysis, the deletion strains were also resistant to lysis induced by thymine starvation of thymine auxotrophs and the loss of viability of these strains after thymine starvation was 100-fold less than in the PBSX parent. The deletion strains had not, however, lost the bacterial autolytic enzymes, since they were still susceptible to lysis when placed under semi-anaerobic conditions.


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