Efficient fluorescence-based localization technique for real-time tracking endophytes route in host-plants...

Area of Science:

• Agricultural Microbiology
• Plant Pathology
• Molecular Biology

Background:

• Biological control agents (BCAs) are crucial for sustainable agriculture, reducing reliance on chemical pesticides.
• Effective tracking of BCAs within host plants is vital for understanding their efficacy and optimizing application strategies.

Purpose of the Study:

• To develop a reliable fluorescence-based technique for labeling and tracking bacterial biological control agents (BCAs) in host-plant interactions.
• To elucidate the localization and colonization routes of BCAs within plants.
• To establish a reproducible method for assessing BCA movement and niches.

Main Methods:

• Transformation of bacterial isolates (PSL, IMC8, PS) with plasmids carrying reporter genes (eGFP, pporRFP).
• Development of antibiotic-resistant competent BCAs.
• Fluorescence microscopy and PCR analysis to confirm BCA localization in Arabidopsis and sorghum.

Main Results:

• High plasmid-mediated transformation efficiencies (up to 84%) were achieved for selected BCAs.
• Fluorescently tagged BCAs were localized to plant roots and hypocotyls, but not leaves or stems.
• Successful confirmation of BCA colonization using fluorescence imaging and PCR.

Conclusions:

• The developed fluorescence-based technique provides high-resolution, reproducible tracking of BCAs within plant tissues.
• This method enables spatial assessment of BCA movement and niches, advancing our understanding of host-plant interactions.
• The protocol facilitates the study of biological and physiological mechanisms underlying plant-microbe interactions.