Live imaging of root–bacteria interactions in a microfluidics setup

Abstract

Plant roots play a dominant role in shaping the rhizosphere, the environment in which interaction with diverse microorganisms occurs. Tracking the dynamics of root-microbe interactions at high spatial resolution is currently limited because of methodological intricacy. Here, we describe a microfluidics-based approach enabling direct imaging of root-bacteria interactions in real time. The microfluidic device, which we termed tracking root interactions system (TRIS), consists of nine independent chambers that can be monitored in parallel. The principal assay reported here monitors behavior of fluorescently labeled as it colonizes the root of within the TRIS device. Our results show a distinct chemotactic behavior of toward a particular root segment, which we identify as the root elongation zone, followed by rapid colonization of that same segment over the first 6 h of root-bacteria interaction. Using dual inoculation experiments, we further show active exclusion of cells from the root surface after colonization, suggesting a possible protection mechanism against root pathogens. Furthermore, we assembled a double-channel TRIS device that allows simultaneous tracking of two root systems in one chamber and performed real-time monitoring of bacterial preference between WT and mutant root genotypes. Thus, the TRIS microfluidics device provides unique insights into the microscale microbial ecology of the complex root microenvironment and is, therefore, likely to enhance the current rate of discoveries in this momentous field of research.

Small molecules below‐ground: the role of specialized metabolites in the rhizosphere

“…Regardless of the great complexity of metabolites secreted by plant roots to the rhizosphere, it is technically difficult to probe the metabolic profiles of native soil exudates in a sensitive, reproducible and comprehensive manner. Even more limited is our capability to examine metabolites in a spatial resolution at varying distances from root systems (Massalha et al ., ). Novel technologies for sampling soil root exudates are thus likely to be employed in the coming years to overcome these limitations.…”

“…Fe deficient conditions Suzuki et al, 2006;Kobayashi andNishizawa, 2012 Mladěnka et al, 2010;Fourcroy et al, 2014;Schmid et al, 2014 Sisó more limited is our capability to examine metabolites in a spatial resolution at varying distances from root systems (Massalha et al, 2017). Novel technologies for sampling soil root exudates are thus likely to be employed in the coming years to overcome these limitations.…”

Recognition of dominant attractants by key chemoreceptors mediates recruitment of plant growth‐promoting rhizobacteria

“…This is the case for symbiotic and nonsymbiotic beneficial bacteria as well as for pathogenic species (Hazelbauer and Lai, ; Hida et al, ; Allard‐Massicotte et al, ; Scharf et al, ; Matilla and Krell, ). Studies of bacterial chemotaxis to plant root exudates, as well as its role in rhizosphere colonization, have received increasing attention in recent years (Scharf et al, ; Massalha et al, ). Amino acids such as Arg, Ala and Ile, as well as organic acids like malic, citric or fumaric acid have been shown to serve as chemoattractants for different rhizobacteria (de Weert et al, ; Gupta Sood, ; Rudrappa et al, ; Tan et al, ; Liu et al, ; Zhang et al, ; Webb et al, ).…”

Dual‐flow‐RootChip reveals local adaptations of roots towards environmental asymmetry at the physiological and genetic levels

“…Over the past years, microfluidic devices have been developed for plant science to facilitate microscopic access to Arabidopsis roots (Meier et al, 2010;Grossmann et al, 2011;Parashar & Pandey, 2011;Busch et al, 2012;Jiang et al, 2014;Massalha et al, 2017), pollen tubes (Horade et al, 2013;Sanati Nezhad et al, 2013) or moss (Bascom et al, 2016). RootChips have been particularly useful for measuring cellular concentration changes of small molecules using genetically encoded nanosensors for nutrients (Grossmann et al, 2011;Lanquar et al, 2014), phytohormones (Jones et al, 2014) or the second messenger calcium (Denninger et al, 2014;Keinath et al, 2015).…”