Isolation, Characterization, and Transfer of Cryptic Gene-Mobilizing Plasmids in the Wheat Rhizosphere

Abstract

A set of self-transmissible plasmids with IncQ plasmid-mobilizing capacity was isolated by triparental exogenous isolation from the wheat rhizosphere with an Escherichia coli IncQ plasmid host and a Ralstonia eutropha recipient. Three plasmids of 38 to 45 kb, denoted pIPO1, pIPO2, and pIPO3, were selected for further study. No selectable traits (antibiotic or heavy-metal resistance) were identified in these plasmids. The plasmids were characterized by replicon typing via PCR and hybridization with replicon-specific probes and other hybridizations. pIPO1 and pIPO3 were similar to each other, whereas pIPO2 was different. None of these plasmids belonged to any known incompatibility group. pIPO2 was selected for further work, and a mini-Tn5-tet transposon was inserted to confer selectability. Plasmid pIPO2 had a broad IncQ plasmid mobilization and self-transfer range among the alpha, beta, and gamma subclasses of theProteobacteria but did not show productive transfer to gram-positive bacteria. Plasmid pIPO2 mobilized IncQ plasmid pIE723 from Pseudomonas fluorescens to diverse indigenous proteobacteria in the rhizosphere of field-grown wheat. Transfer of pIE723 to indigenous bacteria was not observed in the absence of added pIPO2. A specific PCR primer system and a probe were developed for the detection of pIPO2-type plasmids in soil and rhizosphere. Analysis of soil DNA provided evidence for the presence of pIPO2 in inoculated wheat rhizosphere soil in the field study, as well as in the rhizosphere of uninoculated wheat plants growing in soil microcosms. The system failed to identify major reservoirs of pIPO2 in a variety of other soils.

Plant growth promotion abilities and microscale bacterial dynamics in the rhizosphere of Lupin analysed by phytate utilization ability

“…For the same reason, misidentification on the species level is relatively common (LiPuma, 1998). Besides, in the rhizosphere and especially the rhizoplane, horizontal gene transfer occurs frequently (Elsas et al, 1998), as particularly high microbial population densities develop in this region. For example, Burkholderia was shown to be a recipient of large plasmids carrying genes necessary for the ability to degrade the herbicide 2,4-D (Newby et al, 2000).…”

The prevalence and diversity of mobile genetic elements in bacterial communities of different environmental habitats: insights gained from different methodological approaches

“…Mobilising plasmids were also isolated by van Elsas et al [35] when bacterial communities obtained from the rhizosphere of young wheat plants served as donor in triparental matings according to the protocol described by Top et al [34]. Amongst a group of other plasmids, plasmid pIPO2 was isolated in R. eutropha based on its mobilising capacity.…”

“…Thus, to determine the host range of pIPO2, plasmid pIE723 was used as selectable marker for pIPO2 mediated mobilisation. Plasmid pIPO2 was able to mobilise pIE723 as well as to self-transfer to strains belonging to the K, L, and Q subclasses of the proteobacteria [35]. Plasmid pIPO2 mobilised the IncQ plasmid pIE723 from P. £uorescens to indigenous bacteria in the wheat rhizosphere under ¢eld conditions.…”

“…Functioning of the PCR system was shown with the two IncP-9 plasmids pMT2 and pWW0. Primer systems were also designed for the exogenously isolated plasmids pEC10 [22] and pIPO2 [35,37]. Primer systems for the exogenously isolated Hg r plasmid pEC10 were developed based on the sequence of a 300-bp fragment obtained after ampli¢cation by the IncQ oriV system [22].…”

Long‐term manure exposure increases soil bacterial community potential for plasmid uptake

“…Surprisingly, the number of operational taxonomic units (OTUs) was lowest for pIPO2tet (Supporting Information Table S1), the plasmid exhibiting the highest permissiveness. Although the effective phylogenetic host range in the tested soils appears smallest for pIPO2tet, it might exhibit higher transfer frequencies, resulting in a higher total permissiveness, possibly because of its nature as cryptic plasmid (van Elsas et al, 1998). High transfer frequencies are especially important for the maintenance of those plasmids not conferring any beneficial traits to their hosts.…”