Mating experiments were performed at concentrations of each metal that were below the minimum inhibitory concentration (MIC). The definition of ‘transconjugants’ is recipient strains acquired OTC resistance by the mating. The tet(M) gene was detected in transconjugants using PCR (Rahman et al., 2008). Chemical determination of V in Pacific marine sediment was performed. Details of sampling sites and condition are reported in elsewhere (Rahman et al., 2008). Analysis was performed by using inductively coupled plasma-mass spectrometry (ICP-MS) according to the method of Ha et al. (2009). Briefly, the sediment samples were treated with a mixture of HF–HNO3 (1 : 5) and digested by a closed vessel microwave system (Ethos

D, Milestone S.r.l., Sorisole, BG, Italy). The digested solution was heated Rapamycin solubility dmso until acid was removed. The residue was dissolved by HNO3 and diluted with Milli-Q water. Concentrations of 28 trace elements were measured with an ICP-MS (Hewlett-Packard, HP-4500, Avondale, PA). Units of concentrations of trace elements were represented as μg g−1 dry weight. The OTC resistance rate in the sediment has been reported previously (Rahman et al., 2008). The correlation between the V concentration and OTC resistance rate was analyzed in this study.

Susceptibility of all strains used in this ZD1839 research buy study to OTC and various metals was determined as the MIC according to the method described by the National Committee for Clinical Laboratory Standards (NCCLS) (2003). The MICs of OTC and various metals for each strain are shown in Table 1. Exposure to 500 and 1000 μM

V (as VCl3) resulted in a significant increase in the conjugation rate (P < 0.05) (Fig. 1a), and exposure to Ca (CaCl2) also increased the conjugation rate in a dose-dependent manner (Fig. 1b). Exposure of E. coli JM109 to before zinc (Zn as ZnSO4), copper (Cu as CuSO4), and cadmium (Cd as CdCl2) resulted in a decrease in the conjugation rate (Fig. 1c–e). The conjugation rate also increased in an apparent dose-dependent manner upon exposure to mercury (Hg as HgCl2) (Fig. 1f); however, the increase was not significant. It is known that Ca2+ can increase the competency of bacterial cells and induce DNA compaction due to compensation of the DNA electrostatic charge and hydrophobic interactions of the complex sites (Kabanov & Kabanov, 1995), and it is believed that this mechanism contributes to DNA incorporation. Although the mechanism(s) leading to increased rates of OTC resistance following V exposure are not clear, the present study is the first to demonstrate that V can promote conjugation leading to OTC resistance. The MIC of OTC for the recipient E. coli strain was 2 μg mL−1, whereas that of all the transconjugants was significantly higher (256 μg mL−1) (Table 1). The results of PCR analyses indicated that the tet(M) gene was transferred to the recipient E. coli cells, suggesting that the acquisition of OTC resistance occurred through HGT.