, 2000) for auxotrophy. No auxotrophs were detected in our screen of >14 000 clones (∼11 000 of which were recovered following ciprofloxacin treatment as described in Materials and methods). Transposition of phage Mu from its original location to other regions of the chromosome occurs following induction, and is followed by lysis. If the same is true for ECA41, this would explain why the auxotrophy screen failed to yield any colonies: ECA41 may transpose into genes essential for growth on minimal medium, but such cells may not be detected as lysis would follow shortly after. Inward-reading primers flanking the prophages were

designed to detect prophage-less genomes. No ECA41-deficient genomic template was detected, and this is consistent with the stable lysogeny and replicative transposition Daporinad research buy characteristics of Mu. In contrast, an amplicon corresponding to loss of ECA29 was obtained (data not shown). This

amplicon was sequenced, confirming the absence of ECA29 as well as the sequence of the 13-bp direct repeats (GTCAGTAATCGGT) that contribute to the att sites. Navitoclax In contrast with ECA41, excision was precise, reforming the disrupted pflA gene. Spontaneous induction of prophages can result in the presence of phage particles in the culture supernatants of bacterial lysogens. In an attempt to detect these, a filter-sterilized supernatant of a Pa overnight culture was spotted on top agar lawns of 32 different Pa strains, as well as representative strains of E. coli, C. rodentium, Y. enterocolitica and Serratia. No lysis was seen in any case (data not shown). Those Pa strains that carry the prophages are expected to be immune to superinfection by the same phage. Nonetheless, Cobimetinib most of the strains appeared to be naïve to the respective phages (Fig. 1), and could therefore be, in principle, sensitive to infection by these

phages, assuming that the receptor was present. This suggests that no virions were present. Indeed, phage particles were not observed by transmission electron microscopy of culture supernatants following ciprofloxacin treatment (data not shown). It is possible that even though the prophages can excise from the genome, the DNA cannot be packaged into capsids to produce functional virions. Attempts to induce prophage excision and cell lysis using the DNA-damaging agents mitomycin C and UV irradiation were unsuccessful (data not shown). Taken together, these data show that excision of both prophages from the genome occurs, but is a rare, spontaneous and noninducible event, consistent with the relatedness of ECA29 and ECA41 to phage P2 and Mu, respectively. The lysogenic state is stable in these cases, spontaneous induction is rare and they are not inducible by chemical or physical means (Nilsson & Haggard-Ljungquist, 2006; Paolozzi & Ghelardini, 2006).