Natural and Artificial Strategies To Control the Conjugative Transmission of Plasmids

Abstract

Conjugative plasmids are the main carriers of transmissible antibiotic resistance (AbR) genes. For that reason, strategies to control plasmid transmission have been proposed as potential solutions to prevent AbR dissemination. Natural mechanisms that bacteria employ as defense barriers against invading genomes, such as restriction-modification or CRISPR-Cas systems, could be exploited to control conjugation. Besides, conjugative plasmids themselves display mechanisms to minimize their associated burden or to compete with related or unrelated plasmids. Thus, FinOP systems, composed of FinO repressor protein and FinP antisense RNA, aid plasmids to regulate their own transfer; exclusion systems avoid conjugative transfer of related plasmids to the same recipient bacteria; and fertility inhibition systems block transmission of unrelated plasmids from the same donor cell. Artificial strategies have also been designed to control bacterial conjugation. For instance, intrabodies against R388 relaxase expressed in recipient cells inhibit plasmid R388 conjugative transfer; pIII protein of bacteriophage M13 inhibits plasmid F transmission by obstructing conjugative pili; and unsaturated fatty acids prevent transfer of clinically relevant plasmids in different hosts, promoting plasmid extinction in bacterial populations. Overall, a number of exogenous and endogenous factors have an effect on the sophisticated process of bacterial conjugation. This review puts them together in an effort to offer a wide picture and inform research to control plasmid transmission, focusing on Gram-negative bacteria.

FIGURE 1

DNA processing during bacterial conjugation. (1) The relaxase (R) cleaves plasmid DNA at the nic site and forms a covalent intermediate with the 5′ end of the oriT. (2) The T4SS protein machinery recruits the relaxosome through interaction with the T4CP, while the donor DNA is replicated using the uncleaved DNA strand as a template. (3) The relaxase releases the T-strand by a second cleavage reaction at the nic site and acts as pilot protein for the ssDNA to be transferred through the T4SS, helped by the T4CP pumping activity. (4) In the recipient cell, the relaxase carries out the reverse nicking reaction to recircularize the T-strand. (5) The transferred ssDNA is replicated to generate a complete copy of the original plasmid.

FIGURE 2

Natural and artificial mechanisms that control the transmission of conjugative plasmids. Natural mechanisms include RM and CRISPR-Cas systems (encoded by the recipient chromosome), exclusion systems (used to prevent the entrance of related plasmids in the same recipient), and fertility inhibition systems (encoded by plasmids in donor bacteria). Artificial mechanisms interfere with key components of the conjugative process, such as the relaxase, the pilus, or conjugation-related ATPases.

FIGURE 3

Network of interactions between conjugative plasmids that affects their conjugation capacity. Plasmid incompatibility groups are represented by colored circles. Continuous lines show fertility inhibition systems caused by genes in colored rectangles from plasmids in white boxes. Dashed lines show fertility inhibition systems caused by unidentified genes from plasmids in white boxes. See text for further details.

FIGURE 4

Artificial inhibitors of donor-recipient contact (pilus blockers). The tranquilizer chlorpromazine prevents plasmid conjugation and phage infection, possibly by modifying membrane topology. Male-specific bacteriophages bind the pilus tip through their pIII protein, blocking MPF and biofilm formation. Antibodies against conjugative pilus inhibit conjugation of specific plasmids. Zn²⁺ in the mating medium blocks F pilus contact with Zn²⁺-containing receptor sites. Colloidal clay forms a coating on bacterial cells preventing liquid mating, phage infection, and predation. The opioid levallorphan inhibits MPF and adsorption of male-specific bacteriophages, probably by damaging pilus or bacterial membrane. Sodium periodate alters F pili, inhibiting donor fertility and bacteriophage infection. See the section on pilus blockers in the text for additional information.