Biosensors Used for Epifluorescence and Confocal Laser Scanning Microscopies to Study Dickeya and Pectobacterium Virulence and Biocontrol

Abstract

Promoter-probe vectors carrying fluorescent protein-reporter genes are powerful tools used to study microbial ecology, epidemiology, and etiology. In addition, they provide direct visual evidence of molecular interactions related to cell physiology and metabolism. Knowledge and advances carried out thanks to the construction of soft-rot Pectobacteriaceae biosensors, often inoculated in potato Solanum tuberosum, are discussed in this review. Under epifluorescence and confocal laser scanning microscopies, Dickeya and Pectobacterium-tagged strains managed to monitor in situ bacterial viability, microcolony and biofilm formation, and colonization of infected plant organs, as well as disease symptoms, such as cell-wall lysis and their suppression by biocontrol antagonists. The use of dual-colored reporters encoding the first fluorophore expressed from a constitutive promoter as a cell tag, while a second was used as a regulator-based reporter system, was also used to simultaneously visualize bacterial spread and activity. This revealed the chronology of events leading to tuber maceration and quorum-sensing communication, in addition to the disruption of the latter by biocontrol agents. The promising potential of these fluorescent biosensors should make it possible to apprehend other activities, such as subcellular localization of key proteins involved in bacterial virulence in planta, in the near future.

Biocontrol of Biofilm Formation: Jamming of Sessile-Associated Rhizobial Communication by Rhodococcal Quorum-Quenching

“…Confocal microscopy and promoter-probe vectors carrying fluorescent protein-reporter genes are classical tools that have been used to monitor biofilm formation, structure, and development [ 76 , 77 , 78 , 79 , 80 , 81 , 82 ]. They are also powerful tools for localizing and quantifying the AHL-based communication of various plant-associated Gram-negative bacteria [ 57 , 83 , 84 , 85 , 86 , 87 , 88 ]. We used the fine detection capacity of the P. atrosepticum 6276-EI AHL biosensor to show that R. rhizogenes strain 5520 T produces AHLs in the biofilm context ( Figure 3 ).…”