Tn mutagenesis of different fluorescent pseudomonads was achieved by conjugational transfer of the suicide vector pSUP 10141. Pyoverdine negative (Pvd) mutants were detected by the absence of fluorescence on King's B medium and by their inability to grow in the presence of the iron chelator EDDHA [ethylenediamine di(hydroxyphenylacetic acid)]. In ATCC 17400 and three rhizosphere isolates (one and two ), the percentage of Pvdmutants ranged between 0 and 0.54%. In a rhizosphere isolate, this percentage was higher (4%). In these mutants both of the Tn antibiotic resistances (Km and Tc) were stable and the transposon could be detected by hybridization. In Pvdmutants of ATCC 17400, the transposon was found to be inserted twice in the chromosome while single insertions were detected in the DNA of other, randomly tested mutants. In PAO1, where 13.1% of the mutants were Pvd, both antibiotic resistances were rapidly lost and accordingly no transposon insertion could be detected by hybridization. However, the Pvdphenotype was generally stable in these mutants. The plasmid pNK862 containing a mini-Tn transposon was introduced by electroporation into PAO1 and Kmmutants were recovered, 89% of which were Pvdand confirmed to be by PCR amplification of the lipoprotein gene. The mini-Tninsertions were also found to be unstable in PAO1.


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