Visual Evidence of Horizontal Gene Transfer between Plants and Bacteria in the Phytosphere of Transplastomic Tobacco

Abstract

Plant surfaces, colonized by numerous and diverse bacterial species, are often considered hot spots for horizontal gene transfer (HGT) between plants and bacteria. Plant DNA released during the degradation of plant tissues can persist and remain biologically active for significant periods of time, suggesting that soil or plant-associated bacteria could be in direct contact with plant DNA. In addition, nutrients released during the decaying process may provide a copiotrophic environment conducive for opportunistic microbial growth. Using Acinetobacter baylyi strain BD413 and transplastomic tobacco plants harboring the aadA gene as models, the objective of this study was to determine whether specific niches could be shown to foster bacterial growth on intact or decaying plant tissues, to develop a competence state, and to possibly acquire exogenous plant DNA by natural transformation. Visualization of HGT in situ was performed using A. baylyi strain BD413 (rbcL-⌬PaadA::gfp) carrying a promoterless aadA::gfp fusion. Both antibiotic resistance and green fluorescence phenotypes were restored in recombinant bacterial cells after homologous recombination with transgenic plant DNA. Opportunistic growth occurred on decaying plant tissues, and a significant proportion of the bacteria developed a competence state. Quantification of transformants clearly supported the idea that the phytosphere constitutes a hot spot for HGT between plants and bacteria. The nondisruptive approach used to visualize transformants in situ provides new insights into environmental factors influencing HGT for plant tissues.

Transplastomic plants for innovations in agriculture. A review

“…Unfortunately, longer treatments were not tested systematically. Similar results were obtained after visual screening on residues of plant tissue for transformed bacteria expressing an aadA::gfp fusion gene that is restored only upon recombination with total DNA from the same transplastomic plant line studied before (Pontiroli et al 2009). A similar magnitude of gene transfer frequency was also found with DNA purified from a different transplastomic tobacco line and used to transform the same bacterial strain in vitro (De Vries et al 2004).…”

The New Genetics and Natural versus Artificial Genetic Modification

“…However, it is very likely that transformation frequencies are routinely underestimated, as the overwhelming majority of natural bacteria cannot be cultured in the lab. Using methods that restore function to a green fluorescent protein transgene so the transformed bacteria can be seen without the need to culture and select for them, researchers were able to detect transfer of plant DNA to bacteria directly on the surface of intact leaves as well as on rotting, damaged leaves [93,94]. Rotting and damaged leaves release nutrients that promote bacterial growth, and bacteria that can take up foreign DNA are at their most receptive (competent) state for horizontal gene transfer during exponential growth, thus "opportunistic" hotspots for transfer of plant DNA to bacteria are present in plant material infected with pathogens.…”

Genetic Engineering and Sustainable Crop Disease Management: Opportunities for Case-by-Case Decision-Making

“…DNA from residues-transgenic or not-is potentially available for transformation of soil microorganisms via natural processes [136]. Such events are rare but possible [136][137][138][139][140][141]. For prevalent transgenes currently in use, the importance of such a risk is unclear, for several reasons: these genes are already widespread in the environment in their source organisms [137]; crop transgenes sometimes are designed with eukaryotic promotors rather than prokaryotic ones; and they may not be codon-optimized for a recipient microorganism.…”