Determination of the mechanism of retrotransfer by mechanistic mathematical modeling

Abstract

Two mathematical models to elucidate the mechanism of retromobilization (or retrotransfer), that is, the ability of conjugative plasmids to mobilize genes into the cell containing the conjugative plasmid, were developed. This study deals with retromobilization of nonconjugative plasmids (Tra-Mob'). Plasmid transfer was modeled by two mass action models. The first is based on the hypothesis that retromobilization of the TraMob' vector occurs in one step, by means of the pilus formed by the Tra+ plasmid in the original host. In the second model, retromobilization is considered to be a two-step process involving two transfer events. The first step involves the transfer of the Tra+ plasmid from the recipient cell to the donor of the nonconjugative vector, and during the second encounter the nonconjugative vector is mobilized toward the recipient. Since the relationships between the number of transconjugants and the number of recipients for the two models are different, filter matings were performed for short time periods with different initial densities of the recipient population. Comparison of the numbers of transconjugants with the results of the mathematical equations confirmed the hypothesis that retromobilization is a one-step conjugation process.Plasmids belonging to the incompatibility groups IncC, IncJ, IncN, IncP (IncPl), IncQ, and IncW can be maintained in a wide range of bacterial species. Many of these broadhost-range plasmids are able to self-transfer and to mobilize nonconjugative (Tra-) plasmids and sometimes even chromosomal genes into a very wide range of gram-negative bacteria (29). DNA mobilization by IncPl plasmids toward gram-positive bacteria (7, 16, 22, 32), cyanobacteria (11, 34), and yeasts (8) has been demonstrated; the replication range of IncQ plasmids could also be extended to cyanobacteria (11), gram-positive bacteria (6), and plants (1). In addition, IncPl (and some IncN) plasmids mobilize plasmids and chromosomal genes not only in the classical forward direction, i.e., from donor to recipient of the Tra+ plasmid, but also in the reverse direction, i.e., from recipient to donor. This phenomenon of reverse transfer is called retrotransfer and has been observed in both homologous and intergeneric matings.Retrotransfer of chromosomal auxotrophic markers with the IncPl plasmid pULB113 (RP4::Mu3A) was observed in homologous matings with Pseudomonasfluorescens, Salmonella typhimurium, Alcaligenes eutrophus, and Erwinia chrysanthemi, as well as in heterospecific matings between A. eutrophus and Pseudomonas putida carrying pULB113, at frequencies similar to those of the direct mobilization (17). The results obtained in this study gave rise to the assumption that retrotransfer could be an early event in the conjugation process and so would not depend upon the stable acquisition of the conjugative plasmid by the recipient but would occur as a one-step process of bidirectional DNA transfer.Retromobilization of a Tra-Mob-vector, containing the heavy metal resistance genes czc, from Escherichia co...

Retrotransfer or gene capture: a feature of conjugative plasmids, with ecological and evolutionary significance

“…The bidirectional mechanism was associated with the ' one-step ' mathematical model and the unidirectional mechanism with the ' two-step ' mathematical model. The data reported for the RP4 and TOL plasmids fitted the ' one-step ' model (Top et al, 1992 ;Ramos-Gonzales et al, 1994), thus confirming that retrotransfer is an early event. Yet the data of Perkins et al (1994), obtained with exogenously isolated BHR plasmids (Bale et al, 1988), did not completely fit with either model.…”

The complete sequences of plasmids pB2 and pB3 provide evidence for a recent ancestor of the IncP-1β group without any accessory genes

“…Using exactly the same strains, protocols and laboratory (University of Bielefeld) as in the study of Dröge et al (2000), the plasmid transfer frequencies were nearly identical for both plasmids, and comparable to those reported previously for pB2. When the transferability of both plasmids was tested with different strains, using a slightly different filter mating protocol (Top et al, 1992) in a different laboratory (University of Idaho), both plasmids transferred at equally high rates of more than 0?1 per recipient between E. coli strains DH5a and K12Rif (a rifampicin-resistant derivative of MG1655) and between E. coli DH5a and Pseudomonas putida UWC6. Both plasmids transferred at frequencies of approximately 10 23 per recipient from E. coli DH5a to Sinorhizobium meliloti Rm1021 (data not shown) under the same conditions.…”

“…Using exactly the same strains, protocols and laboratory (University of Bielefeld) as in the study of Dröge et al (2000), the plasmid transfer frequencies were nearly identical for both plasmids, and comparable to those reported previously for pB2. When the transferability of both plasmids was tested with different strains, using a slightly different filter mating protocol (Top et al, 1992) recipient from E. coli DH5a to Sinorhizobium meliloti Rm1021 (data not shown) under the same conditions. This lack of difference in transferability between the two plasmids is in agreement with their very high sequence similarity.…”

Modelling the spatial dynamics of plasmid transfer and persistence

“…Plasmid transfer experiments for which quantitative results are reported were performed using 0.45 mm pore, 25 mm diameter filters (Whatman) on LB agar plates (15 g agar l 21 ) essentially as described before (Top et al, 1992). Liquid cultures of donor and recipient strains were grown overnight at 30 uC in LB broth (with Tc 10 mg ml 21 for the donor), and centrifuged and resuspended in an equal volume of LB broth.…”