THE REGION OF A BACTEROIDES CONJUGAL CHROMOSOMAL TETRACYCLINE RESISTANCE ELEMENT WHICH IS RESPONSIBLE FOR PRODUCTION OF PLASMID-LIKE FORMS FROM UNLINKED CHROMOSOMAL DNA MIGHT ALSO BE INVOLVED IN TRANSFER OF THE ELEMENT

Large (> 50 kilobases) conjugal chromosomal tetracycline resistance (Tc(r)) elements have been found in many human colonic Bacteroides strains. Recently, N.B. Shoemaker and A.A. Salyers (J. Bacteriol. 170:1651-1657, 1988) reported that some of these Tc(r) elements appeared to mediate production of plasmidlike forms, NBU1 and NBU2, from an unlinked region of the chromosome of Bacteroides uniformis 0061. Production of the plasmidlike forms and the transfer frequency of the Tc(r) elements were both enhanced by preexposure to tetracycline. Thus it appeared that genes involved in production of plasmidlike forms (Plf activity) might be coregulated with transfer genes and that Plf activity might have a role in transfer of the Tc(r) elements. By screening subclones of a Tc(r) element, Tc(r) Em(r) DOT, we have shown that the genes necessary for Plf activity on the Tc(r) element are within a 10-kilobase region adjacent to the Tc(r) gene. Subclones of this region were then used to construct insertional gene disruptions in a Tc(r) element, Tc(r) ERL, which is closely related to the Tc(r) Em(r) DOT element. Two of the disruption mutants were Plf-. Both had reduced transfer frequencies, one (ΩRDB2) 102-fold lower than that of the wild-type element and the other (ΩRDBT) 104-fold lower. ΩRDB2 was also deficient in the ability to mobilize coresident plasmids, whereas ΩRDBT exhibited nearly wild-type mobilization activity. The phenotypes of the mutants indicate that there are at least two genes necessary for Plf activity and that both may be involved in transfer of the element. The third disruption mutant (ΩRDB1), which expressed Plf constitutively, also had a transfer frequency 102-fold lower than that of the wild-type element and was deficient in mobilization of coresident plasmids. The relationship between Plf genes and transfer, therefore, appears to be a complex one.