Effect of mercury addition on plasmid incidence and gene mobilizing capacity in bulk soil

Abstract

Two methods of exogenous plasmid isolation were used to evaluate and compare the plasmid incidence and gene mobilizing capacity of eight soils before and after the addition of mercury. Biparental matings (BM) were performed using Pseudomonas putida UWC1 as recipient to obtain conjugative mercury resistance (HgR) plasmids. To obtain mobilizing plasmids, the transfer of the IncQ vector pMOL187 from Escherichia coli into Ralstonia eutropha was selected in triparental matings (TM). The numbers of donor, helper and recipient cells used in the matings were kept the same for each soil sample, which provided a way to compare plasmid incidence and gene mobilizing capacity on the basis of the numbers of transconjugants obtained. Using BM prior to mercury addition, plasmids that confer HgR were obtained only from 1 of 8 soils. However, following the addition of mercury to these soils, HgR transconjugants were obtained from 5 of the 8 soils, and among these broad (BHR) and narrow-host-range (NHR) plasmids were found at variable ratios. Similarly, these same five soils were the only ones to yield mobilizing plasmids by using TM; all of which were found to confer HgR and to be BHR plasmids. This suggests that HgR plasmids were the main cause of the increased mobilizing capacity observed in the mercury treated soils. Notably, there were three soils in which no plasmids that confer mercury resistance were obtained although the soils had at least 105 HgR cfu g soil−1, indicating that resistance to mercury may not be encoded on plasmids that could be transferred to the recipients used. Plasmids from 177 transconjugants obtained by BM and TM could be classified into 20 groups based on size and restriction fragment patterns, and none belonged to known classes of BHR plasmids. This indicates that there are diverse replicons capable of mediating the dissemination of mercury resistance.