Transfer of Tn925 and plasmids between Bacillus subtilis and alkaliphilic Bacillus firmus OF4 during Tn925-mediated conjugation

Guffanti, Arthur A.; Quirk, Philip G.; Krulwich, Terry A.

doi:10.1128/jb.173.5.1686-1689.1991

Cited by 7 publications

(3 citation statements)

References 19 publications

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“…However, these observations differ somewhat from those of Guffanti et al (16), who recently reported a relatively high frequency of plasmid mobilization between B. subtilis and Bacillus firmus by the conjugative transposon Tn925. In addition, Torres et al (33) reported a high level (e.g., 50% or more) of cotransfer of chromosomal markers with a cis-located Tn925 in both B. subtilis and E. faecalis mating systems.…”

Section: Discussioncontrasting

confidence: 56%

Conjugative transfer of Tn916 in Enterococcus faecalis: trans activation of homologous transposons

Flannagan

Clewell

1991

J Bacteriol

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tet. Experiments showed that the presence of one element (e.g., Tn916AE) in a recipient cell did not prevent the uptake of the other (e.g., Tn916) (7, 24), indicating an absence of negatively acting factors relating to entry exclusion or incompatibility. The presence of a transposon on the recipient chromosome also failed to prevent the zygotically induced excision-insertion that occurs at a high frequency when Tn916 enters passively on a conjugative plasmid (7). The data argue against the presence of a strong, trans-acting negative regulator being produced by an established element.In the present study we focus on the interaction that occurs between Tn916 and Tn9J6AE when they reside together in a donor cell. Evidence is presented showing that the transfer of one element results in a high-frequency trans activation of the other. Comobilization of resident plasmids was also detected, but at frequencies much lower than that involving the transposon. MATERIALS AND METHODSBacterial strains, plasmids, and media. The bacterial strains and plasmids used in this study are listed in Tables 1 and 2, respectively. Cultures of E. faecalis were grown in Todd-Hewitt broth (THB; Difco) at 37°C. Escherichia coli cultures were grown at 37°C in LB medium (11) for preparation of competent cells and in THB for plasmid isolation. Antibiotics, when present, were used in the following concentrations except as noted: tetracycline, 10 ,ig/ml (to maintain pAM420 in E. coli, a concentration of 60 p.g/ml was used); erythromycin, 10 ,ug/ml; chloramphenicol, 25 ,ug/ml; fusidic acid, 25 p.g/ml; streptomycin, 1,000 jig/ml; spectinomycin, 500 ,ug/ml; rifampin, 25 ,ug/ml. Rifampin was from Calbiochem, whereas all other antibiotics were from Sigma.DNA analyses. Chromosomal DNA was prepared by a standard protocol (35). Plasmid DNA was purified by an alkaline lysis procedure modified from that of Ish-Horowicz 7136 JOURNAL

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Section: Discussioncontrasting

confidence: 56%

Conjugative transfer of Tn916 in Enterococcus faecalis: trans activation of homologous transposons

Flannagan

Clewell

1991

J Bacteriol

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“…Shaded nucleotides correspond to the promoter that has been mapped for the cadmium resistance determinant of p1258 (29). Arrows indicate an 11-bp inverted repeat analogous to the region of dyad symmetry that has been implicated in the regulation of S. aureus cad gene expression (29 (8). 2501 GTTCTCTTGCTCTTAGATACACAGGMAATCCCCCTMTCCMCCCGMGGTTTMTTTTTTTCTAMGCTCGATACMCACTGATTTACTMCACCCGT Antiporter activity of everted membrane vesicles from E. coli NM81 transformed with either pJB22 or the pGEM7Zf(+) control plasmid.…”

Section: Methodsmentioning

confidence: 99%

The cadC gene product of alkaliphilic Bacillus firmus OF4 partially restores Na+ resistance to an Escherichia coli strain lacking an Na+/H+ antiporter (NhaA)

et al. 1992

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A 5.6-kb fragment of alkaliphilic Bacilus firmus OF4 DNA was isolated by screening a library of total genomic DNA constructed in pGEM3Zf(+) for clones that reversed the Na+ sensitivity of Escherichia coli NM81, in which the gene encoding an Na+/H+ antiporter (NhaA) is deleted (E. Padan, N. Maisler, D. Taglicht, R. Karpel, and S. Schuldiner, J. Biol. Chem. 264:20297-20302, 1989 Alkaliphilic Bacillus species growing at a pH higher than 10 maintain a cytoplasmic pH that is much more acidic than that of the external medium by using an Na+ cycle consisting of Na+/H+ antiporters and a group of Na+/solute symporters (11,12). The antiporters catalyze the electrogenic exchange of extracellular H+ for intracellular Na+, driven by the proton motive force established by respiration (12,14). Solute uptake coupled to Na+ entry completes the cycle, which allows cells growing at pH 10.5 to maintain an intracellular pH of approximately 8.3 (7).The alkaliphile Na+ cycle has been extensively characterized physiologically, and our recent efforts are aimed at identifying and characterizing the genes encoding the antiporter activity. We have used a screen based on the observation of Padan et al. (21) alkaliphile gene distinct from nhaC into its genome by recombination with a plasmid from the alkaliphile DNA library (10).In this report, we describe a third transformant that confers Na+ resistance to E. coli NM81 without supplying a structural gene for an Na+/H+ antiporter. Rather, Na+ resistance is associated with the expression of a small protein normally involved in the efflux system bestowing cadmium resistance (33). MATERIALS AND METHODSBacterial strains and growth conditions. Alkaliphilic Bacillus firnus OF4 was grown routinely at pH 10.5 or 7.5 on highly buffered malate-containing media at 30°C as described previously (7). Tests of the cadmium resistance of B. firmus OF4 811M required the use of a modified medium to avoid precipitation of the cadmium. The basal medium for growth at pH 7.5 or 9.0 consisted of 50 mM Tris-HCl, 0.1% (NH4)2SO4, and 0.1 mM MgSO4. Cadmium resistance at pH 10.2 was monitored in a basal medium containing 50 mM sodium CAPS [3-(cyclohexylamino)-1-propanesulfonic acid] buffer instead of Tris-HCl. All media were supplemented with 50 mM DL-malate, 0.1% yeast extract, and 50 ,ug of methionine per ml. E. coli NM81 (21) was grown at 37°C in LBK medium (21) containing kanamycin (50 ,ug/ml). Transformants were selected on LBK-kanamycin medium plus ampicillin (100 ,ug/ml), and 0.4 to 0.7 M NaCl was added to screen for Na+-resistant transformants. E. coli JM109 (2) and DH5atMCR (GIBCO-BRL) were used for routine cloning procedures; transformants were selected and maintained on SOB medium (2) containing ampicillin. 4878on June 7, 2019 by guest

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“…Tn916 and Tn925 (very similar to Tn916 in structure and function; Christie et al 1987) have been shown to have a broad host range and are capable of conducting cross-species transfer (Gawron-Burke and Clewell 1984;Christie et al 1987;Bertram et al 1991). These transposons have been used to move plasmid DNA between Bacillus species (Naglich and Andrews 1988b;Guffanti et al 1991) and as shuttle vehicles (Norgren et al 1989;Torres et al 1991).…”

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confidence: 99%

Conjugative transposition of Tn916 and Tn925 in Bacillus popilliae

Dingman¹

1999

Can. J. Microbiol.

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Interspecies transfer of the conjugative transposons Tn916 and Tn925 into B. popilliae Pj1 occurred using Enterococcus faecalis and Bacillus subtilis CU4049 as transposon donors. Tn916 was stably maintained in B. popilliae Pj1 following growth without selective pressure and was successfully introduced into the plasmid-containing B. popilliae strains NRRL B-2524, Ch1, and KLN4 using E. faecalis CG110. In B. popilliae, expression of the tetracycline resistant determinants on Tn916 and Tn925 provided resistance to 25 µg/mL and 50 µg/mL tetracycline, respectively. An erythromycin resistant determinant, present in Tn916∆E, was also functional in B. popilliae Pj1 and provided resistance to 1 mg/mL erythromycin. Transfer of Tn916 into E. faecalis, B. subtilis, and between B. popilliae strains was accomplished using a transposon-containing strain of B. popilliae as donor. Efforts to transfer Tn916 between E. coli and B. popilliae were unsuccessful.

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Transfer of Tn925 and plasmids between Bacillus subtilis and alkaliphilic Bacillus firmus OF4 during Tn925-mediated conjugation

Cited by 7 publications

References 19 publications

Conjugative transfer of Tn916 in Enterococcus faecalis: trans activation of homologous transposons

Conjugative transfer of Tn916 in Enterococcus faecalis: trans activation of homologous transposons

The cadC gene product of alkaliphilic Bacillus firmus OF4 partially restores Na+ resistance to an Escherichia coli strain lacking an Na+/H+ antiporter (NhaA)

Conjugative transposition of Tn916 and Tn925 in Bacillus popilliae

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