Development and applications of a DNA labeling method with magnetic nanoparticles to study the role of horizontal gene transfer events between bacteria in soil pollutant bioremediation processes

Abstract

Horizontal gene transfers are critical mechanisms of bacterial evolution and adaptation that are involved to a significant level in the degradation of toxic molecules such as xenobiotic pesticides. However, understanding how these mechanisms are regulated in situ and how they could be used by man to increase the degradation potential of soil microbes is compromised by conceptual and technical limitations. This includes the physical and chemical complexity and heterogeneity in such environments leading to an extreme bacterial taxonomical diversity and a strong redundancy of genes and functions. In addition, more than 99 % of soil bacteria fail to develop colonies in vitro, and even new DNA-based investigation methods (metagenomics) are not specific and sensitive enough to consider lysis recalcitrant bacteria and those belonging to the rare biosphere. The objective of the ANR funded project “Emergent” was to develop a new culture independent approach to monitor gene transfer among soil bacteria by labeling plasmid DNA with magnetic nanoparticles in order to specifically capture and isolate recombinant cells using magnetic microfluidic devices. We showed the feasibility of the approach by using electrotransformation to transform a suspension of Escherichia coli cells with biotin-functionalized plasmid DNA molecules linked to streptavidin-coated superparamagnetic nanoparticles. Our results have demonstrated that magnetically labeled cells could be specifically retained on micromagnets integrated in a microfluidic channel and that an efficient selective separation can be achieved with the microfluidic device. Altogether, the project offers a promising alternative to traditional culture-based approaches for deciphering the extent of horizontal gene transfer events mediated by electro or natural genetic transformation mechanisms in complex environments such as soil.

Zooming in to acquire micro-reaction: Application of microfluidics on soil microbiome

“…Burmeister et al (2018) found that P. putida was not able to produce yellow fluorescence when bacteria were cocultured in a space-separated device, while contact between the cells caused a change in the fluorescent signal from red to yellow, indicating the occurrence of HGT. Pivetal et al (2015) designed a new microfluidic system composed of micromagnets to monitor HGT. Magnetic nanoparticles were used to label plasmid DNA molecules and magnetic microfluidic devices were then employed to capture and separate the recombinant cells with nanoparticle signals.…”

Magnetic Nylon 6 Nanocomposites for the Microextraction of Nucleic Acids from Biological Samples

“…Solid-phase protocols allow more efficient NA isolation than other methods [16,18,28]. Magnetic solid-phase extraction (MSPE) is a fast and easy procedure with high NA yield and purity [18,26,[31][32][33][34][35][36][37][38][39][40][41]. MSPE may be an automated method for NA microextraction without specific equipment [17,18].…”

Innovative technique for patterning Nd–Fe–B arrays and development of a microfluidic device with high trapping efficiency

“…Later on, in 1996, a relatively more compact device had been developed for sorting and trapping human peripheral lymphocytes tagged with ferromagnetic particles [47]. However, because of the aforementioned reasons on the need for small-scale devices (Lab-on-chip) alongside the advances in synthesis techniques like lithography, researchers' focus shifted towards micro-and millimeter-size permanent magnets [22,[48][49][50][51][52][53][54][55][56][57][58][59][60]. Moreover, reducing the size of the magnets increases the magnitude of the magnetic field gradient [43], ergo, more magnetic particles could be separated/trapped.…”